Blut, 1977 Jan, 34(1), 49 - 52 A mofidied method for measuring the filtrability of erythrocytes; Imre S; A modified method based on Teitel's original procedure for measuring the filtrability of erythrocytes was described . The filter paper was not placed into a funnel but to the exit at the bottom of a 2 ml glass column . The filtration of the red cell suspension was quantitated by measuring the decrease of the volume in the column. J Immunol, 1977 Jan, 118(1), 146 - 50 Immunologic reactivity of the lung . III . Effects of corticosteroids on alveolar macrophage cytotoxic effector cell function; Hunninghake GW et al.; The effects of various in vitro and in vivo regimens of corticosteroid administration on guinea pig alveolar macrophages were studied . Corticosteroid-induced immunosuppression was assessed by the effect of drug administration on the total numbers and functional capabilities of alveolar macrophages as measured by the PHA-induced and antibody-dependent cellular cytotoxicity assays against sheep red blood cell targets . In vivo administration of either hydrocortisone sodium succinate (100 mg/kg/one dose) or cortisone acetate (100 mg/subcutaneously for 7 days) caused a marked increase in the numbers of alveolar macrophages recovered from the teased lung cell suspensions and 4 and 24 hr, respectively, after the last injection . Both regimens of corticosteroid administration cause similar levels of peripheral blood lymphocytopenia and monocytopenia, 4 and 24 hr, respectively, after the final injection . Neither in vitro hydrocortisone (0, 1, and 10 mug/ml), nor hydrocortisone (100 mg/kg), in vivo had any effect on either the PHA-induced or antibody-dependent cellular cytotoxicity of alveolar macrophages . In marked contrast, cortisone acetate, depo-preparation which gives sustained elevations of plasma cortisol levels similar to those found for a brief period after i.v . injection of hydrocortisone caused a marked decrease in cytotoxic effect on function of alveolar macrophages suspensions . In a separate experiment, the suppressed killer cell function of the alveolar macrophages from steroid-treated animals was found not to be related to an intrinsic defect in killing of bound target cells since the defect in killing could be overcome by increasing the density of antibody and PHA on the target cells. Neuroendocrinology, 1977, 24(3-4), 232 - 40 Acute decrease of blood thyroid hormone by isovolemic exchange transfusion as a stimulus for pulse TSH release and its modifications by CNS Lesions; Langer P et al.; The level of thyroid hormone in blood has been acutely decreased by about 20--30% within 20 min with the aid of isovolemic exchange transfusion (IET) of thyroid hormone free blood cell suspension (THFBCS) in three groups of rats: (1) intact control (C); (2) median eminence (ME)-lesioned; (3) thalamus (TH)-lesioned . The levels of thyroxine (T4) and TSH were measured by specific radioimmunoassay (RIA) before the transfusion and for 180 min after that . The level of T4 increased to the initial value at about 120 min after IET in C and ME-lesioned rats, while in TH it remained decreased until 180 min . Furthermore, at the end of IET the level of TSH in C and ME-lesioned rats was about 2--3 time higher than the initial value and its continuous decrease was observed until 180 min . In contrast, in TH-lesioned rats the increase of TSH level was delayed, being significant at 60 min only . It was concluded that IET of THFBCS acts as a stimulus of acute TSH release which was remarkably inhibited in TH-lesioned animals . In addition, the destruction of most of the ME did not apparently influence this response, as shown by essentially similar data obtained in C and ME-lesioned rats. Scand J Immunol, 1977, 6(12), 1251 - 61 Antibody-dependent cytotoxicity mediated by cells eluted from synovial tissues of patients with rheumatoid arthritis and juvenile rheumatoid arthritis; Abrahamsen TG et al.; Cell suspensions containing an average of 78% lymphocytes were obtained from synovial tissues of 26 patients with rheumatoid arthritis and 10 patients with juvenile rheumatoid arthritis . These cells were shown to mediate cytotoxicity against 51Cr-labeled chicken erythrocytes sensitized with a rabbit anti-chicken erythrocyte antiserum . Nylon column filtration of the cells increased the proportion of lymphocytes and usually also the cytotoxicity, which suggested that at least some of the effector cells were lymphocytes . The cytotoxic activity of the cells obtained from rheumatoid synovial tissue was always lower than that obtained with the patients' peripheral blood lymphocytes . No significant change in cytotoxicity of normal peripheral blood lymphocytes was observed after these cells had been treated in the same manner as the rheumatoid synovial tissues. Am J Hematol, 1977, 3, 237 - 44 The use of filtration techniques for the lysis and study of red blood cells; Hultquist DE et al.; A filtration technique for the gentle lysis of erythrocytes has been developed using cellulose triacetate membranes . When cell suspensions are filtered under nitrogen pressure, lysis occurs at the surface of the filter in such a way that the cell ghosts are retained on the filter . The contents of the cell are extruded through the pores of the filter without mixing with the cell suspension . Cell ghosts and intact erythrocytes have been collected on membranes and examined by electron microscopy . These preparations have the advantage of being free of the structural artifacts that result from centrifugation . In addition, the filter facilitates preparation for electron microscopy by providing a support for the sample during fixation and then dissolving during the dehydration of the sample. Acta Biol Med Ger, 1977, 36(3-4), 543 - 8 {Viability, ATP and 2,3-DPG content of resuspended erythrocytes during several-week storage in cold}; Strauss D et al.; Leukocyte-and thrombocyte-poor packed red cells obtained from ACD or . ACD-AG blood were resuspended to a hematocrit of about 55% and stored at 4 degrees C . The resuspension solution consisted of xylitol, inorganic phosphate, bicarbonate, adenine (A) and guanosine (G) solved in water . In one case glucose, citrate and sucrose were also added, in another one, sorbitol . The 2,3-DPG and the ATP level remained for a longer period in the sorbitol-xylitol-medium than in the glucose-xylitol-medium . The ATP content in the red cell suspension was higher than in packed cells . Higher ATP values were obtained in red blood cells from whole blood with adenine and guanosine . The survival rate of resuspended red blood cells in glucose-AG-citrate-sucrose medium was about 80--85% after 3 weeks of storage and 77% after 6 weeks with a higher range. Folia Biol (Praha), 1977, 23(4), 235 - 42 Action of A RNase and AS RNase on growth of cells in vitro; Cinatl J et al.; The effect of bovine pancreatic A RNase and bull seminal vesicle AS RNase on proliferation of HeLa cells and LEP cells in vitro was studied . The ribonucleases were used in doses of 0.1, 1.0, 10.0, and 100.0 microgram per ml of medium and 100 000 inoculated cells . Proliferation was evaluated by the growth curves . Both in single and long-term experiments . A RNase in all doses used had no effect on cell proliferation, whereas AS RNase exerted inhibitory activity beginning with the dose of 10 microgram, and affected more markedly the LEP cells (a diploid non-malignant cell line) than the malignant heteroploid HeLa cells . When added to the cell suspension, AS RNase was shown by indirect immunofluorescence to bind to HeLa cell membranes in all phases of growth, whereas AS RNase was found on membranes of LEP cells only when they were derived from the log phase of growth. Am J Pathol, 1977 Jan, 86(1), 123 - 31 Glucose-6-phosphate dehydrogenase isoenzymes in acinar epithelial cells separated from the mammary gland of the rat at intervals during the course of lactation; Scalise MM et al.; Acinar epithelial cells were purifed from suspensions of cells from the lactating mammary glands of rats . As described previously, this purififaction was accomplished by sedimentation in an isokinetic gradient of Ficoll (polysucrose) in tissue culture medium, and the purity of the separated cells was established by electron microscopy and by histochemical markers . Isoenzymes of glucose-6-phosphate dehydrogenase were investigated at various intervals during lactation in separated populations of stromal and acinar cells . Acinar cells contained three- to eightfold more glucose-6-phosphate dehydrogenase activity than did stromal cells . The proportions of the respective isoenzymes varied during the course of lactation, and the observed changes were parallel in purified acinar cells and in the lactating mammary glands from which the cell suspensions were obtained . The availability of purified acinar cells in the investigation of interactions between hormones and cells from the mammary gland permits a greater degree of specificity than has been possible in the study of mammary cell suspensions which contain myoepithelial cells, duct cells, acinar cells, endothelial cells, fibroblasts, fat cells, mast cells, plasma cells, and blood cells. Biull Eksp Biol Med, 1977 Jan, 83(1), 73 - 5 {Changes in the immunologis status of the body during carcinogenesis detected by the physico-chemical characteristics of cells of the immunocompetent system}; Stepanova EN et al.; The state of cell population of the lymph nodes and the thymus was studied on a model of rat sarcoma of the hip(induced by dimethlbenzanthracene) by the method of fluorescent sounds . Statistically significant changes of incorporation of the negatively charged sound 1-anilino-8-naphthalinosulfonate (ANS) into cell suspension, depending on the stage of sarcoma development, were detected . The appearance of cell forms with a lesser hydrophobic character of the surface and with less binding points for the ANS were noted at the early carcinogenesis periods . It is supposed that these cells were referred to immature ones . A conclusion was drawn on the applicability of the method of fluorescent sound for recording changes in the immunological state of the organism in carcinogenesis. Microbios, 1977, 19(75), 17 - 26 Effect of phenoxyethanol on the permeability of Escherichia coli NCTC 5933 to inorganic ions; Gilbert P et al.; Concentrations of phenoxyethanol which retarded the growth rate of Escherichia coli NCTC 5933 in nutrient broth, stimulated the rates of respiration and total oxygen uptake of cell suspensions with glucose as carbon source, and were able to dissipate artificially induced membrane proton gradients . In cells with repressed oxidative phosphorylating activity, no stimulation of respiration was observed . These actions were characteristic of uncouples of oxidative phosphorylation . Similar concentrations of the drug caused additional increased permeability of the cytoplasmic membrane to K+ but not to Li+, Na+, Ca++, Mg++, NO-3, Cl-, So--4, or PO---4 . Drug induced permeability of the membrane to protons and potassium ions were not found to be directly coupled. Scand J Immunol, 1977, 6(12), 1305 - 15 Studies of antibody-dependent cytotoxicity in a plaque assay: evidence of human monocyte-like effector cells; Thorsteinsson L et al.; Two different methods for evaluating 'in vitro' cytotoxicity against antibody-coated target cells mediated by mononuclear leukocytes were compared . One was a plaque assay for identification of the cytotoxic cell and the other the classical chromium release assay for antibody-dependent cytotoxicity (ADCC) . A marked decrease in plaque-forming cells (PFC) was observed in a cell suspension depleted of peroxidase-positive cells and cells with membrane-bound Ig (B lymphocytes) by fractionation on a nylon fiber column . In contrast, the ADCC activity was considerably increased by these depletions . A similar effect was obtained by removal of phagocytic cells with iron . These results, together with the observations after depletion of E-RFC (T lymphocytes) or EA-RFC (Fc-receptor-bearing cells), suggest that the PFC in the assay system used were of monocytic origin and differnet from the cells responsible for the ADCC. Int Arch Allergy Appl Immunol, 1977, 55(1-6), 131 - 4 Role of T and adherent cells in in vitro response of nude mouse spleen cells to bacterial lipopolysaccharide; Takigawa M et al.; Thy 1.2-positive cells and adherent cells were depleted from nude mouse spleen cell suspensions by treatment with anti-Thy 1.2 antisera plus complement and by passage through Sephadex G-10 column, respectively . The results of 3H-TdR uptake and appearance of IgM containing cells induced in vitro by lipopolysaccharide (LPS) were similar in both untreated and depleted spleen cells . Our findings confirm T and adherent cell independence of LPS response in nude mice. J Immunol Methods, 1977, 17(3-4), 329 - 36 A direct hemolytic plaque assay on polylysine-coated coverslips for scanning electron microscopy of antibody forming cells; Pan S et al.; A direct immunoplaque assay was utilized without agar or other semi-solid matrix for examining antibody secreting cells by scanning electron microscopy . For this purpose a polylysine procedure was used whereby 'charged' coverslips absorbed onto their surface an even monolayer of target sheep erythrocytes . Hemolysin secreting splenocytes derived either from mice immunized in vivo with SRBC or from normal or primed splenocytes immunized in vitro with the same antigen formed large numbers of hemolytic plaques on the RBC-coated polylysine treated coverslips . SEM examination of such plaque-forming cells revealed the presence of cells with the typical surface morphology of lymphocytes present in whole spleen cell suspensions . Many of the lymphocytes were covered with numerous microvilli while a small percentage were relatively free of such villi and exhibited a much smoother surface . Examination of lymphocytes secreting hemolytic antibody on polylysine coated coverslips sensitized with target erythrocytes thus permits the direct examination and analysis of surface morphologic features and also permits a direct comparison with other features revealed by light and fluorescent microscopy. J Pathol, 1977 Jan, 121(1), 1 - 8 Characteristics of mononuclear cell populations in chronically inflamed synovial membranes; Meijer CJ et al.; Mononuclear cells infiltrating synovial membranes in chronic synovitis were characterised both in situ and in cell suspensions by surface markers and histochemical techniques . T-lymphocytes were the predominant infiltrating cell in rheumatoid arthritis as well as in other forms of chronic arthritis, including ankylosing spondylitis and arthritis associated with Crohn's disease . B-lymphocytes were found exclusively in rheumatoid synovial membranes . These cells were demonstrable both in true germinal centres and, focally and diffusely, in nodular mononuclear infiltrates lacking the histochemical characteristics of germinal centres . The synovial lining cells, unlike mononuclear phagocytes, had no demonstrable receptors for C3 and Fc. Int Arch Allergy Appl Immunol, 1977, 54(4), 364 - 73 Experiments on the passive sensitization of human basophils, using quantitative immunofluorescence microscopy; Stallman PJ et al.; In previous experiments a correlation was found between the amount of IgE on human basophils and the IgE serum level . The results of these experiments are shown to fit a free exchange model with an approximately constant K-value . We investigated whether the free IgE-receptors on the basophils, which should be present according to this model, could be saturated by incubating the cell suspensions with IgE myeloma protein or with sera from patients with an elevated IgE serum level . Various incubation conditions were applied in sensitizing the leukocytes, and cells from both atopic and nonatopic subjects were used for testing, but no increase in basophil-bound IgE could be measured with the quantitative immunofluorescence technique . Nor could we demonstrate a significant dissociation of cell-bound IgE in a period of 24 h . Therefore another hypothesis is put forward, in which receptor turnover is taken into account: we assumed that that free receptors on the basophils had a shorter functional half-life, than the half-life of the IgE-receptor complexes . This model would explain the contradictory results mentioned above. Microsc Acta, 1977 Jan, 79(1), 43 - 5 The use of porous plastic filters in the preparation of cell suspensions; Pearce FL; Suspensions of cells were prepared by dissociation of various tissues with the proteolytic enzyme pronase . Debris, undisrupted tissue and cell clumps were effectively removed from the suspensions by passage through VYON F porous plastic filters whereas isolated cells were recovered without significant losses . The viability of the cells, as judged by dye exclusion and subsequent behavior in tissue culture, was not affected by the treatment. Bull Cancer, 1977, 64(2), 225 - 40 Morphological and functional differentiation and classification of cutaneous lymphomas; Burg G et al.; It was the purpose of our studies to achieve reevaluation of the histo- and cytomorphology of cutaneous lymphomas by means of additional enzymecyto-chemical and funtional tests . Skin biopsy specimens from 99 patients with cutaneous lymphomas and pseudolymphomas were stained for HE, Giemsa, PAS, Gormori, several hydrolytic enzymes and peroxidase . In cell suspensions extracted from skin lesions B and T cell differentiation was performed using surface markers . In addition tissue cells were tested for PHA response in suspensions and for intracytoplasmatic Ig on smears . Based on these studies cutaneous lymphomas were filed, within the "Kiel" classification of low grade and high grade malignant lymphomas according to their histological, enzymecytochemical and immunological features . It got evident that B cell and T cell lymphomas of low grade malignancy in the skin both present distinct histological patterns, indicating areas B and T cell microenvironmental specificity. J Immunol Methods, 1977, 16(1), 1 - 13 Optimal conditions for the separation of rat T lymphocytes on anti-immunoglobulin--immunoglobulin affinity columns; Kondorosi E et al.; The separation of rat T lymphocytes was investigated on anti-Ig--Ig columns . A simple and efficient method for the purification of rat Ig by precipitation of rat serum with sodium sulfate is presented . Protein binding characteristics of glass and plastic beads, as solid support of affinity columns, are described, as well as optimal parameters for coating beads with rat Ig (with BSA, ribonuclease and lysozyme, as comparison) . Binding of Ig was primarily dependent on the concentration of the Ig solution . Maximal strong binding of Ig (6.2 X 10(3) molecules per micron2 of bead surface) was reached a 400 microng per ml concentration of purified Ig solution during 20 min of incubation . Higher concentrations increased only the amount of loosely bound Ig on the surface of beads whereas the amount of firmly bound Ig remained unchanged . Fractionation of lymphoid cell suspensions on anti-Ig--Ig affinity columns prepared at optimal conditions resulted in highly purified T-cell suspensions containing less than 1% of lymphocytes bearing surface Ig receptors. Ann Immunol (Paris), 1977 Jan-Mar, 128(1-2), 47 - 9 {Cellular interaction in the pokeweed-mitogen induced terminal differentiation to polyclonal immunoglobulin production in human B lymphocytes in vitro}; Robert M et al.; The differentiation of human B lymphocytes in vitro into Ig secreting cells, as measured by a radioimmunoassay rendered specific to IgG and IgM, in response to a polyclonal stimulant, requires the participation of several cellular types . Upon poleweed-mitogen stimulation, B-cell enriched and monocyte containing cell suspensions prepared by elution of nylon wool adherent cells, when mixed with various concentrations of T cells purified by filtration through nylon wool column and depletion of EAC-rosette forming cells, synthesize 5 to 20 times more Ig than non separated lymphocytes . These experiments which demonstrate the cooperation of certain T cells also suggest that other cells eliminated by nylon wool adherence act as suppressor cells . The cooperation of these T cells is independent of their proliferation and can be obtained with allogeneic T cell. Transfusion, 1977 Jan-Feb, 17(1), 16 - 22 A method for radioactive antiglobulin testing with 125I labeled anti-IgG; Jenkins DE Jr et al.; The present report describes a method for 125I anti-IgG antiglobulin testing . The method involves the use of a labeled DEAE cellulose IgG fraction of goat anti-human IgG globulin . Leukocyte-free red blood cells were used in the test procedure . Both the labeled antiserum and the red blood cell suspensions were diluted in normal rabbit serum to prevent non-specific uptake of labeled antiserum by normal, uncoated red blood cells . The results of the study show that the method is highly reproducible and relatively easy to perform once the labeled anti-IgG is prepared . Furthermore, the use of labeled antiserum permits analysis of antiserum specificity by radioimmunoelectrophoresis . The problem of determining anti-IgG/IgG binding ratios remains the major disadvantage to using the method to determine precise amounts of human IgG coating red blood cells. Physiol Chem Phys, 1977, 9(6), 533 - 8 Effect of haemagglutinating virus of Japan on the potassium compartmentation and the membrane potential of human erythrocytes; Utsumi K et al.; HVJ induces release of potassium and changes in membrane potential of certain types of cells . These changes are quite dependent on incubation temperature and are inhibited by the addition of con A to cell suspensions prior to the addition of viruses. Endocr Res Commun, 1977, 4(2), 159 - 69 An improved bioassay for ACTH determination; Liotta AS et al.; An improved method for the bioassay of ACTH has been developed using short-term culture of adrenal cell suspensions derived from intact rats . Six separate pools of adrenals were used and cells derived from the same pool were compared after acute dispersion and overnight culture . The ED50, defined as the concentration of ACTH required to produce half maximal steroidogenesis, for cultured cells was 35.4+/-2 pg/ml compared to 240+/-60 pg/ml for cells used immediately after dispersion . The minimum concentrations of ACTH necessary to produce a response significantly above basal levels were 0.98+/-.10 pg/ml and 12.1+/-3 pg/ml respectively . Response characteristics of the cultured cells were highly reproducible . The incubation volume employed in this study was 0.25 ml, so the ED50 expressed as dose of ACTH per tube was actually 8.8 pg for the cultured cells . Such sensitivity provides requisite methodology for the measurement of ACTH under varying physiological conditions. Prostaglandins, 1977 Jan, 13(1), 25 - 31 Cyclic AMP response to prostaglandin E1 in mononuclear cells from peripheral blood and synovial fluid of patients with rheumatoid arthritis; Zurier RB et al.; The cyclic AMP response to prostaglandin E1 (PGE1) was studied in peripheral blood (PB) and synovial fluid (SF) mononuclear cells from patients with rheumatoid arthritis (RA) . The PGE1 induced accumulation of cyclic AMP was consistently (7 of 8 patients) less in cell suspensions derived from SF than in suspensions of equivalent numbers of mononuclear cells obtained simultaneously from PB . The high PB/SF cyclic AMP ratio was seen most clearly at the lowest concentration (10(-6)M) of PGE1 tested . There was no correlation between the patients' therapy and cyclic AMP response to PGE1 . The high PB/SF cyclic AMP ratio was not accounted for by the presence of platelets in PB cell suspensions. Transplantation, 1977 Jan, 23(1), 53 - 9 Mixed heart cell-lymphocyte reactions and graft survivals in Ag-B-compatible rats; Kashiwabara H et al.; A method of dissociation of the rat heart cell for the mixed lymphocyte culture purpose is described . The treatment of the young rat heart with 0.1% collagenase-hyaluronidase solution yielded satisfactory heart cell suspension . The hear cells, thus obtained after mitomycin C treatment but not irradiation, stimulated allogeneic lymphocytes in the presence of 2-mercaptoethanol . In the Fischer to Lewis combination compatible at the Ag-B locus, strong reactions of Lewis lymphocytes to the dissociated heart and skin cells of the Fischer rat were related to the acute rejection of heart and skin grafts, while negative reactions by the kidney and spleen cells reflected a prolonged survival of the kidney graft . The role of skin- and hear-specific antigens in the rejection phenomenon is discussed. J Bioeng, 1977 Jan, 1(2), 61 - 77 The preferential adsorption of hemoglobin to polyethylene; Horbett TA et al.; Hemoglobin adsorption to foreign surfaces has not previously been considered in studies of blood-material interactions, despite the fact that hemoglobin is the most abundant protein present in blood . A hemoglobin-like protein was detected on a number of surfaces exposed to blood plasma, serum, and red cell suspensions . Hemoglobin adsorption to polyethylene from plasma was found to approximately equal the amount of adsorption of albumin and fibrinogen . The high relative affinity of hemoglobin for polyethylene was further confirmed by adsorption isotherm and direct competition experiments . The data from all four experimental methods support the following ranking of plasma protein affinity for polyethylene: Hemoglobin greater than fibrinogen greater than albumin congruent to gamma-globulin. Vox Sang, 1977, 33(5), 257 - 65 Preparation and specificity testing of a rabbit anti-human thymocyte serum (with 1 colour plate); Brutel de la Riviere A et al.; The preparation of a specific anti-T cell serum, applicable in the indirect immunofluorescence technique on cell suspensions, smears of peripheral blood and bone marrow and on tissue sections is described . Rabbits were immunized with thymocytes; after removal of antibodies against species-specific antigens and antigens common to leucocytes by absorptions with red cells and granulocytes, specificity for thymocytes was obtained by repeated absorptions with CLL cells, used as B cell equivalents . Subsequently, the IgG fraction of the absorbed antiserum was isolated . This antibody preparation was tested with various types of blood cells as well as with cell suspensions depleted or enriched in T cells . For the study of tissue sections it had to be absorbed with liver powder . When studying lymphatic tissue it was found to stain only thymus-dependent areas in the specimens tested. J Immunol Methods, 1977, 15(3), 291 - 7 Temperature dependence of antigen-specific rosette formation by lymphocytes from immunised mice; Wansbrough-Jones M et al.; Spleen cell suspensions from mice undergoing a secondary response to sheep erythrocytes (SRBC) contained about one tenth as many specific antigen-binding, rosette-forming cells (RFC) when they had been washed at 37 degrees C instead of 4 degrees C before rosetting . This difference was correlated with the presence of IgG anti-SRBC antibody in the serum, and the 37 degrees C washings of immunised spleen cells could passively allergise non-immune spleen cells at 4 degrees C for specific rosette formation which was inhibitable by anti-mouse F(ab)2 serum . The RFC from actively immunised mice were lymphocytes and not macrophages by morphological and cytochemical criteria . It is suggested that the 37 degrees C-labile RFC are lymphocytes to which IgG antibodies bind in the cold . These data indicate that in the use of antigen-binding cell assays to monitor immunological responses, it is necessary to wash lymphocytes at 37 degrees C before testing. Ann Immunol (Paris), 1977 Jan-Mar, 128(1-2), 137 - 9 {H-2 specificity and auto-reactivity of autologous rosette-forming cells from adult thymectomized mice spleen cells}; Charreire J et al.; We have previously shown that the number of autologous rosette-forming cells (A-RFC) found in adult thymectomized mice (A-Tx) spleens was 15 times higher than that found in spleens of intact controls . We have therefore investigated both the specificity of A-RFC and their relationship with autoreactivity . Immunoadsorption experiments showed that while the incubation on allogeneic monolayers did not bring any modification in A-RFC levels . This result suggests that the formation of autologous rosettes is the manifestation of a true recognition event associated with the H-2 complex . A-Tx mice spleen cells injected into syngeneic recipients induced an enlargement of the afferent popliteal lymph node (LN) . Ficoll-Triosil A-RFC depleted A-Tx spleen cells lost their capacity to induce this popliteal proliferation . Conversely, A-RFC recovered spleen cell suspensions obtained from the pellet of the Ficoll Triosil gradient kept their capacity of inducing auto-reactivity . Thus, it seems that A-RFC and lymphocytes stimulating the afferent LN belong to the same sub-population. J Immunol Methods, 1977, 14(3-4), 267 - 9 Rapid identification of monocytes in a mixed mononuclear cell preparation; Tucker SB et al.; A method for identifying monocytes by the "non-specific esterase" stain is described . This method is particularly applicable to mononuclear cell suspensions obtained by Ficoll--Hypaque density gradient separations and allows rapid as well as accurate determinations. Dev Biol Stand, 1976 Dec 13-15, 37, 235 - 40 The testing of cell invasiveness in embryonic tissue and in cell culture; Ikic D et al.; The subcutis of 11-day-old chick embryos, 15- to 18-day-old mouse embryos and the skin muscle tissue of human fetus were stretched on lens paper in a Petri dish and seeded with a piece of LED-Widr or HDC cell sheet . Mem + 10% FCS was added . After three days incubation the material was fixed, sectioned and H-E stained . In the second part of our investigation several cell lines were prepared on cover slips . When confluent, the cultures were inoculated with 1-2 drops of cell suspension of another cell line . Three days after the seeding of cell suspension on the confluent cell sheet the cultures were fixed and stained . It can be seen in the organ cultures that LED-Widr cells closely adhere to the embryonic tissue which in some places they are invading . In tissue culture examination we found that heteroploid cells seeded on a confluent sheet of diploid cells adhered but that a diploid cell seeded on epithelial-like heteroploid cells would not adhere . The testing of invasiveness in embryonic tissue in organ culture showed that it can be a useful model . By seeding suspended cells of fibroblast lines on a confluent sheet of heteroploid cell lines their contact inhibition or their invasive ability can be determined . The seeding of heteroploid cell lines on diploid cell lines can be used as a model for the study of the mechanism of invasion into normal tissue and for the in vitro study of invasion inhibiting properties of certain substances. Aust J Biol Sci, 1976 Dec, 29(5-6), 443 - 51 Preparation of metabolically active cell suspensions from wool roots; Ward KA; A method is described for the preparation of metabolically active cell suspensions from plucked wool roots using the proteolytic enzyme trypsin . The suspensions consist of a heterogeneous population of cells which appear similar in morphology to follicle bulb cells, differentiating keratinocytes and possibly cells of the inner root sheath . The concentrations of trypsin and of inorganic ions for optimum activity of the suspensions have been determined, and the inclusion of EGTA was found to increase the yield of cells . The cell suspensions incorporate {14C}leucine, {3H}uridine and {3H}thymidine into acid-insoluble products, and are sensitive to the action of cycloheximide and emetine, but not to chloramphenicol or rifampin . Autoradiography has shown that the cells believed to be derived from the follicle bulbs show the greatest activity. J Cell Sci, 1976 Dec, 22(3), 657 - 70 Assay of intercellular adhesiveness using cell-coated Sephadex beads as collecting particles; Vosbeck K et al.; A simple, rapid and precise method, based on a previous method, for measuring relative rates of intercellular adhesion is described . DEAE-Sephadex beads were treated with nitrocellulose in order to allow cells to grow on their surfaces . Balb/c 3T3 and Balb/c 3T12 cells were used to characterize the assay . They formed confluent cell layers on nitrocellulose-treated DEAE-Sephadex . These cell-coated beads were employed to collect 32P-labelled cells from single cell suspensions . Since they formed statistically uniform, large collecting surfaces, the collection of labelled cells was markedly improved as compared to the original assay . The cell-coated beads collected a large percentage of the labelled cells in a short time . The percentage of cells collected was independent of the concentration of labelled cells in the assay mixture, and the collection was linear for approximately 60 min . The variability between replicate assays was usually +/- 5% . The assay allows the rapid and precise determination of intercellular adhesion in large numbers of individual samples . These features make it useful to screen for effects of different treatments on intercellular adhesions. Arch Microbiol, 1976 Dec 1, 111(1-2), 137 - 44 Growth of Hansenula polymorpha in a methanol-limited chemostat . Physiological responses due to the involvement of methanol oxidase as a key enzyme in methanol metabolism; van Dijken JP et al.; Hansenula polymorpha has been grown in a methanol-limited continuous culture at a variety of dilution rates . Cell suspensions of the yeast grown at a dilution rate of 0.16 h-1 showed a maximal capacity to oxidize excess methanol (QmaxO2) which was 1.6 times higher than the rate required to sustain the growth rate (QO2) . When the dilution rate was decreased to 0.03 h-1, QmaxO2 of cells increased to a value of more than 20 times that of QO2 . The enzymatic basis for this tremendous overcapacity for the oxidation of excess methanol at low growth rates was found to be the methanol oxidase content of the cells . The level of this enzyme increased from 7% to approximately 20% of the soluble protein when the growth rate was decreased from 0.16 to 0.03 h-1 . These results were explained on the basis of the poor affinity of methanol oxidase for its substrates . Methanol oxidase purified from Hansenula polymorpha showed an apparent Km for methanol of 1.3 mM in air saturated reaction mixtures and the apparent Km of the enzyme for oxygen was 0.4 mM at a methanol concentration of 100 mM . The involvement of an oxygen dependent methanol oxidase in the dissimilation of methanol in Hansenula polymprpha was also reflected in the growth yield of the organism . The maximal yield of the yeast was found to be low (0.38 g cells/g methanol) . This was not due to a very high maintenance energy requirement which was estimated to be 17 mg methanol/g cells X h. Arch Microbiol, 1976 Dec 1, 111(1-2), 111 - 5 Induction of D-6-hydroxynicotine oxidase in resting cells of Arthrobacter oxidans; Reeves HC et al.; Resting cell suspensions of Arthrobacter oxidans were shown to synthesize the inducible enantiozyme, D-6-hydroxynicotine oxidase, in the presence of D-nicotine or D-6-hydroxynicotine . The corresponding L-enantiomers, as well as gamma-methylaminopropyl-(6-OH-pyridyl-3)-ketone, which is the product of the reaction catalyzed by the enzyme, were ineffective as inducers . L-6-Hydroxynicotine inhibited induction by D-nicotine and D-6-hydroxynicotine while L-nicotine inhibited induction by D-6-hydroxynicotine and had no effect on induction by D-nicotine . Enzyme induction was also found to be inhibited by glucose, 2-deoxy-D-glucose and by several intermediates of the tricarboxylic acid cycle . An absolute requirement for protein synthesis and for oxygen was also demonstrated to be necessary for the reactions involved in the covalent attachment of flavin adenine dinucleotide to pre-existing precursor protein to yield the catalytically active D-6-hydroxynicotine oxidase. Br J Cancer, 1976 Dec, 34(6), 619 - 25 Stimulation of autologous blood lymphocytes by malignant lymphoma cells and homogenates; Ludgate ME et al.; The blastogenic response to autologous blood lymphocytes to whole-cell suspensions and to homogenates obtained from malignant lymphoma tissue has been investigated . Spleens were obtained from patients in whom laparotomy was performed for staging of malignant lymphoma . Cell suspensions prepared from tumour nodules were treated with mitomycin C and allowed to react with separated autologous blood lymphocytes for 6 days . Lymphocyte stimulation was measured by liquid scintillation counting after exposure to 3H-TdR . Cultures were also prepared in which autologous lymphocytes were treated with spleen tumour homogenate . Control experiments used spleens from staging procedures in which no tumour deposits were present, and normal spleens removed incidentally during other operations . In the controls, the uptake of TdR was never more than twice that of unstimulated lymphocytes . Greater degrees of lymphocyte stimulation were seen in 6 out of 14 patients, using whole tumour cells, and in 7 out of 16 patients, using tumour homogenates . The results indicate an antigenic difference between tumour and host cells, and suggest that lymphocytes can react to a tumour-associated antigen. Lab Invest, 1976 Dec, 35(6), 550 - 7 Tissue factor in cultured cells: pharmacologic effects; Maynard JR et al.; Tissue factor activity in suspension cultures of WISH amnion cells is modulated by pharmacologic doses of agents which alter membrane structure and function . Lysosomal stabilizing steroids (hydrocortisone, dexamethasone, aldosterone, prednisolone, and estradiol) suppress the change in activity which follows subculture; lytic steroids (testosterone and progesterone) are ineffective . Chloroquine both increases the specific activity and extends the time before return to the basal level . Dimethyl sulfoxide and ouabain suppress the complete expression of activity but do not inhibit the subsequent decay . The effect of cytochalasin B is complex, the drug being either suppressive or slightly stimulatory depending on the time of addition . Cyclic nucleotides (AMP or GMP) or insulin do not regulate the expression of tissue factor in these cells . A dramatic increment and prolongation of activity occurs when colchicine or vinblastine is added to the cell suspension shortly after subculture; there is much less stimulation by griseofulvin . Lumicolchicine has no effect while deuterium oxide is inhibitory . From these experiments, we conclude that increased membrane fluidity or altered secretory processes resulting from microtubule disruption stabilize tissue factor in cultured cells . Since contradictory results were obtained with agents which stabilize lysosomes or inhibit transport, the role of these cellular functions in tissue factor production or decay is unclear. J Cell Biol, 1976 Dec, 71(3), 921 - 32 Effects of glutathione-oxidizing agents on microtubule assembly and microtubule-dependent surface properties of human neutrophils; Oliver JM et al.; In human peripheral blood polymorphonuclear leukocytes and lymphocytes, GSH-oxidizing agents promote the movement of surface-bound concanavalin A (Con A) into caps and inhibit the assembly of microtubules (MT) that is normally induced by Con A binding . Con A capping and inhibition of MT assembly occur when GSH levels in cell suspensions are decreased by 30-70%, and return to GSH to control levels is accompanied by the appearance of cytoplasmic MT and by inhibition of the capping response with Con A . Oxidation of GSH markedly stimulates the hexose monophosphate shunt, and regeneration of GSH occurs rapidly . The data indicate that MT cannot be assembled or maintained in the face of decreased GSH levels . Thus, GSH homeostasis becomes critical during physiological events such as phagocytosis which simultaneously induce the assembly of MT and the production of agents like H2O2 that can oxidize GSH. Ann Med Interne (Paris), 1976 Dec, 127(12), 865 - 72 {Morphologic criteria of surface studies by electron microscopy and classification of lymphoproliferative syndromes . Critical study}; Reyes F et al.; Surface associated immunoglobulins (s.Ig) have been detected on human lymphocytes, in normal individuals and in disease, by an immunoelectron microscopic method using peroxidase-labeled antibodies . Experiments have been carried out on fixed cell suspensions, in order to avoid membrane alterations induced by anti-immunoglobulin antibodies . Normal human blood B lymphocytes have a villous surface . However this relationship between microvilli and detectable s.Ig, as found in the normal state, is not confirmed by examinating various T and B cell proliferative states . Thus surface morphology alone is not sufficient for classifying cells in disease . The precise nature of mononuclear cells from hairy cell leukemia remains nuclear. Arch Microbiol, 1976 Dec 1, 111(1-2), 73 - 6 Spectroscopic properties and related functions of the stigma measured in living cells of Euglena gracilis; Benedetti PA et al.; The spectroscopic properties of stigma inside green and dark-grown cells and of isolated stigma globules have been studied by means of a microspectrophotometer build in the Laboratory . On the base of these results and of the analysis of the absorption spectra of a stigma suspension, cell suspension and the cell methanolic extracts, it can be inferred that pigments localized in the stigma are free carotenoids which are not closely packed and do not show an ordered arrangement . Furthermore, the efficiency of the stigma as shading device is duscussed. Am Rev Respir Dis, 1976 Dec, 114(6), 1099 - 105 Characterization of immunocompetent cells recovered from the respiratory tract and tracheobronchial lymph node of normal guinea pigs; Gorenberg DJ et al.; To gain insight into the structure of the lung's immune system, the identity and function of immunocompetent cells in the broncho-alveolar air spaces and tracheobronchial lymph nodes of normal guinea pigs were examined . Guinea pig thymus-derived (T) lymphocytes (mediators of cellular immunity) were identified by their ability to form rosettes with rabbit erythrocytes (E rosettes) . Bone marrow-derived (B) lymphocytes (mediators of humoral immunity) were identified by their ability to form rosettes with sheep erythrocytes sensitized with antibody and complement (EAC rosettes) and by the presence of surface immunoglobulin, detected by direct immunofluorescence . Total lung lavage yielded 14+/-5.0 X 10(6) (mean +/- SD) cells . There were 68+/-5 per cent cells of the monocyte-macrophage series as judged by morphology, the ingestion of latex particles, and the uptake of neutral red; 20+/-9 per cent were eosinophils, and 12+/-5 per cent were lymphocytes . Stable populations of T and B lymphocytes were recovered from guinea pig airways and tracheobronchial lymph nodes . T cells represented 76 per cent of lymphocytes from the airways and 64 per cent of lymphocytes in tracheobronchial lymph node; B cells equalled 14 per cent and 30 per cent, respectively . The functional potential of T cells present in the lung and tracheobronchial lymph node was demonstrated by their proliferative response to phytohemagglutinin . These results indicated that, although macrophages are the predominant cell type, viable T and B lymphocytes are present in the guinea pig broncho-alveolar air space . The composition of these cells is comparable to what has been observed in normal human lungs and provides preliminary evidence that the guinea pig may be a useful model for humans . This report also describes a method using 2 vital stains whereby macrophages, lymphocytes, and rosetted lymphocytes can be distinguished in unseparated cell suspensions prepared from bronchial aspirates. Acta Pathol Microbiol Scand {C}, 1976 Dec, 84C(6), 489 - 94 Freezing of rat lymphocytes . III . Freezing of plaque-forming cells and restoration by frozen-thawed normal cells of antibody production in irradiated rats; Hem E; The present investigation is an extension of earlier work with dimethyl sulphoxide (DMSO) protected frozen-thawed rat lymphocytes . In the present work it is shown that some 85-90% of the haemolytic plaque-forming cells (PFC) survived the freeze-thaw process . Irradiated rats were restored with fresh and frozen-thawed cells and immunized against sheep red blood cells (SRBC) . Evidence is presented that a restrictive control of the PFC response by suppressor cells present in the spleen cell suspension is lost during the freeze-thaw process, giving a higher number of PFC/spleen in recipients of frozen-thawed mixed spleen and lymph node cells than in rats receiving the corresponding fresh preparations. Cancer, 1976 Dec, 38(6), 2296 - 309 The immune response at the tumor site in lung carcinoma; Ioachim HL et al.; The local immune response to lung cancer was investigated by histologic and immunologic means . Distinctive patterns of stromal cellular reaction, characteristic for different histologic types of lung carcinoma, were recognized . The amount of cellular infiltration was highest in squamous cell carcinomas and lowest or nonexistent in oat cell carcinomas . Within the various histologic categories the well-differentiated tumors appeared to be accompanied by more reactive cells than the poorly differentiated ones; there was no relation between tumor necrosis and cellular infiltration . The plasma cells were distinctly associated with squamous cell carcinomas; their number in the stroma was proportionate to the degree of differentiation and the presence of keratin produced by the tumors . Eluates with a high content of immunoglobulins were recovered from pleural effusions and from solid lung carcinomas by dissociation of antigen-antibody complexes . These preparations reacted positively in indirect immunofluorescence tests with tissue cultures and with fresh suspensions of lung carcinoma cells, but not with tissue culture cells of most nonpulmonary tumors or with cell suspensions of normal adult and fetal lung . Similarly prepared fractions of noncarcinomatous pleural effusions did not react with lung cancer cells. Immunology, 1976 Dec, 31(6), 943 - 51 Analysis of immunosuppression generated by the graft-versus-host reaction . II . Characterization of the suppression cell and its mechanism of action; Shand FL; Spleen cells from (CBA X C57/BL) F1 mice undergoing graft-versus-host (GVH) reaction induced by injection of parental cells 7-14 days previously are capable of suppressing an immune response by normal or primed F1 spleen cells to chicken erythrocytes and levan in vivo and sheep erythrocytes in vitro . The cells in these GVH spleens which were responsible for the suppression were sensitive to treatment with anti-0 serum, resistant to 900 rad irradiation in vivo and not retained by anti-immunoglobulin columns . Suppressor activity in vitro was present only in the non-adherent fraction of these GVH cell suspensions . Furthermore, the T-cell fraction, purified by affinity chromatography, suppressed the in vitro response of macrophage-depleted normal F1 cells to DNP-levan . Collectively, these observations imply that suppressor T cells generated by GVH reaction can affect B-cell functions directly without intermediary macrophage participation . Spleen cells from (CBA X C57/BL) F1 mice undergoing GVH reaction induced by C57/BL cells were depleted of their F1 content by treatment with anti-CBA alloantiserum . The suppressive activity of the residual donor component was still expressed against other F1 cells (AKR X C57/BL) which were H-2 compatible with the original host, but not against H-2-incompatible cells (DBA/1 X C57/BL) F1 . However, the latter were suppressed in the presence of (CBA X C57/BL) F1 cells . Thus, interaction of donor T cells with F1 target cells containing those H-2 antigens towards which they are sensitized is mandatory for the subsequent manifestation of immunosuppressive activity . GVH cells suppressed the response of primed F1 cells in double Marbrook chambers when the two populations were separated were by a cell-impermeable membrane, provided the GVH suspension contained F1 cells to which donor T cells were sensitized . This suggests that soluble factors are involved in the mechanism of GVH-induced immunosuppression. Int J Cancer, 1976 Nov 15, 18(5), 687 - 96 Agglutination reactions of spontaneous canine tumour cells, induced by concanavalin A, demonstrated by an isotopic assay; Betton GR; Quantitative assessment of the agglutination of 51Cr labelled canine cell suspensions to canine kidney cell monolayers has been performed over a range of concanavalin A concentrations . Agglutination was observed with all cell cultures tested, comprising four spontaneous canine melanomas, two canine mammary carcinomas, a benign mammary tumour and a contact-inhibited kidney cell line . The melanomas tested showed strong specific inhibition of concanavalin A agglutination by 10(-2)M alpha-methyl-D-glucopyranoside . Inhibition of agglutination of mammary tumour and kidney cells was weaker and less specific . Agglutination was inhibited at 4degrees C . Reduced agglutination to glutaraldehyde-fixed mono-layers was observed in the case of mammary tumours but was absent when contact-inhibited kidney cells were tested . The specificity of the reaction for transformed cells and the parameters involved are discussed. J Natl Cancer Inst, 1976 Nov, 57(5), 1157 - 67 An assessment of intratumor phagocytic and surface marker-bearing cells in a series of autochthonous and early passaged chemically induced murine sarcomas; Pross HF et al.; Single cell suspensions from five different 3-methylcholanthrene-induced tumors in CBA mice were examined in the autochthonous host and sequentially for 5-11 passages . They were also examined for Fc receptor-bearing, phagocytic, theta antigen-positive, and surface immunoglobulin-bearing cells . The preparations contained a high proportion of phagocytic and marker-bearing cells both in the original host and during early passage . This proportion was consistent for any particular tumor and passage . Between different tumors, however, the proportions were sufficiently different to allow the tumor to be identified on this basis; this suggested that various chemically induced tumors may be unique in their tumor-host relationship as measured by the type of cells which infiltrate them . With on-going early passage of the tumors, the proportion of marker-bearing cells decreased to a constant level in most instances, mainly because of a reduction in the percentage of phagocytic cells . The tumor with the least macrophages (MBQA, less than 5%) consistently appeared more rapidly and killed the host more rapidly than did the tumor with the most macrophages (MBQD, 15-30%), but was not significantly different in its growth rate . The theta antigen- and Fc receptor-positive cells within these tumors were derived from the animal receiving the tumor inoculum, and thus represented host cell infiltration of the tumor . The results were discussed with reference to fundamental concepts of the immunology of chemically induced tumors and the importance of host cell infiltration within these tumors. J Natl Cancer Inst, 1976 Nov, 57(5), 995 - 1007 Characterization of mammotrophs separated from the human pituitary gland; Hymer WC et al.; Human pituitary tissues from 27 patients and 7 persons post mortem were dissociated into single cell suspensions . On the average, 23% of the cells were mammotrophs . The concentration of prolactin in these suspensions averaged 3.8 ng/1,000 cells . After cell separation by velocity sedimentation at unit gravity, mammotrophs and other cell types were enriched twofold to threefold . The separated mammotrophs retained structural integrity at light and electron microscopic levels . In eight separation experiments, cells recovered from different gradient regions were assayed for intracellular prolactin levels . In cells from "normal" subjects, 8.5% of the prolactin recovered from the gradient was associated with large mammotrophs, whereas in patients with breast cancer, 28% of the hormone was associated with large mammotrophs . The number of mammotrophs recovered from this gradient region (beyond fraction 6) was doubled in breast cancer (2 expts) . These mammotrophs showed areas of hypertrophied Golgi and endoplasmic reticulum . Culture of the separated cells from 1 patients with diabetes and 2 patients with breast cancer for 21 days showed that mammotrophs in the upper gradient fractions (diabetic) secreted seven times more hormone than those in the lower regions, whereas those mammotrophs from patients with breast cancer that fell to the lower gradient regions secreted 15 times more prolactin than did those in the upper regions . These data suggest that pituitaries of patients with breast cancer contain a small pool (10-20%) of hypertrophied mammotrophs that have the potential for significant secretory activity in vitro. J Reprod Fertil, 1976 Nov, 48(2), 347 - 53 A highly efficient method for washing mammalian spermatozoa; Harrison RA; A simple method has been developed for washing spermatozoa: the cell suspension is layered over a solution containing Ficoll and, after centrifugation, the original suspending medium remains as an undiluted layer above the Ficoll solution while the spermatozoa form a loose pellet at the bottom . Removal of the supernatant layers is easily and completely accomplished by aspiration . When ram spermatozoa were washed by this method, as much as 99-6% of the original medium could be removed during a single washing cycle, while up to 98% of the cells were recovered . Mechanical damage to the cells was minimal and repeatability was high. Blut, 1976 Nov, 33(5), 301 - 12 Re-entry of resting leukaemic blood cells into proliferation in human acute leukaemia during diffusion chamber culture; Hoelzer D et al.; From 17 patients with different forms of acute leukaemia, mononuclear blood cells were cultured in diffusion chambers (DC) implanted intraperitoneally into pre-irradiated mice . In 14 patients, growth of blast cells could be observed during the culture period of up to 21 days . To question whether this growth of blast cells was due only to proliferation of the initially proliferating fraction or whether a re-entry of resting leukaemic cells into proliferation was involved, various 3H-thymidine (3H-TdR) labelling studies were carried out . The absolute increase of blast cells in EC showed no correlation with the fraction of leukaemic blast cells in DNA-synthesis in the implanted cell suspension as measured by 3H-TdR labelling in vitro . Furthermore, in 2 patients where the kinetic behaviour of initially labelled leukaemic blast cells was followed during DC culture, the increase in total blast cells could only be attributed to a small extent to proliferation of those cells initially in the cell cycle . Lastly, "in vivo" labelling during the culture period showed that in one case 25% and in another case 60% of the blast cells in DC were proliferating . The conclusion is that, owing to the stimulation of the diffusion chamber milieu and possibly also due to removal of an in vivo inhibition, in most cases of acute leukaemia resting leukaemic blast cells can apparently re-enter the active cell cycle . This has relevance for an understanding of the self-maintenance of the leukaemic cell population and may also be a reason for relapse of leukaemia after the usual cytostatic drug treatment which affects mainly the proliferating leukaemic cells. Am J Clin Pathol, 1976 Nov, 66(5), 883 - 91 An evaluation of the Haema-Count MK-40 blood counting system; Weisbrot IM et al.; The Haema-Count MK-40, a semiautomated blood counting system, determines hemoglobin, erthrocyte count, leukocyte count, and hematocrit values of blood cell suspensions prepared with a small automatic pipetter-diluter . It is similar to the MK-3, which the authors previously evaluated, but has automatic coincidence correlation and a modified prime and rinse cycle . Its performance was compared with those of standard methods, i.e., the single-channel Coulter Counter and manual cyanmethemoglobin and microhematocrit methods . Precisions for hemoglobin determinations and for leukocyte and erythrocyte counts were equal to those of the reference (comparative) methods . Patients comparisons for those determinations had only small intermethod variability and small clinically insignificant biases . The hematocrit channel was the least precise . With a modified method of calibration, the authors obtained patient comparisons without statistically significant bias from the microhematocrit . Calibration was stable for all channels during the course of the study. Am J Anat, 1976 Nov, 147(3), 357 - 73 Dissociation of epithelial cells from rabbit trachea and small intestine with demonstration of APUD endocrine cells; Sonstegard KS et al.; In this study the entire epithelial lining of tracheas and a 15-cm segments of small intestine were dissociated into individual cell components after 45-minute incubation with 1% pronase . Light and electron microscopy of isolated cells confirmed good morphologic preservation of various epithelial cell types dissociated from the trachea and small intestinal mucosa . Of particular interest was the recovery and preservation of APUD endocrine cells, which are known to be widely dispersed amongst various non-endocrine epithelial cells in both the trachea and small intestine . The APUD cells were demonstrated in dissociated cell preparations by a formaldehyde-induced fluorescence method, Grimelius' silver nitrate stain, and electron microscopy . The isolated APUD cells retained their characteristic features, e.g., amine-handling properties, argyrophilia and cytoplasmic dense-core vesicles . The cell dissociation method described in this report provides high yields of viable epithelial cells in single cell suspensions which are suitable for further cell separation into homogeneous populations of single kinds of cells, including the APUD endocrine cells . Availability of methods for isolation of tracheal and intestinal APUD cells will facilitate further studies, in vitro, on secretory, metabolic and functional aspects of these cells. Blood, 1976 Nov, 48(5), 743 - 53 Size distribution, electronic recognition, and counting of human blood monocytes; Loos H et al.; During a study on the separation of human blood monocytes from lymphocytes, a method was developed to recognize and count monocytes by electronic means . Lightscattering (Cytograf, Bio/Physics), and changes in electrical resistance (Channelyzer, Coulter) were used to size mononuclear leukocytes directly in cell suspensions . Both methods revealed a size distribution profile in which two populations of mononuclear leukocytes could be distinguished . The largest cells were virtually eliminated after phagocytosis of iron particles . We confirmed that these cells were monocytes by three different criteria: the intracellular lysozyme activity, the number of phagocytes, and the percentage of cells with kidney-shaped nuclei . The highly significant correlations we found showed that monocytes could be recognized and counted by electronic sizing . For this method, purified mononuclear leukocyte preparations had to be used, since the presence of erythrocytes, platelets, and polymorphonuclear cells interfered . Statistical analysis revealed that electronic sizing permitted discrimination of differences in monocyte content of 4.5%, with a probability of 95% . It was calculated that this sensitivity of electronic monocyte counting was about three times higher than the sensitivity of microscopic methods . Since 100,000 cells can be sized within a few seconds, not only the efficiency of the preparation but also minor changes in the size of monocytes and lymphocytes introduced during the isolation can be followed. Mikrobiologiia, 1976 Nov-Dec, 45(6), 941 - 5 {Hydrogen production by the cyanobacterium Anabaena variablis in the light}; Gogotov IN et al.; Light of low intensity (less than or equal to 25-10(5) erg-cm(-2)-sec(-1)) stimulates hydrogen production by cell suspensions of Anabaena variabilis in the presence of glucose, pyruvate or formate . The maximum rate of hydrogen production in the presence of these substrates was observed at light intensities of 650, 1400 and 2250 erg-cm(-2)-sec(-1), respectively . The rate of oxygen production by the cells increases while the rate of hydrogen evolution decreases with increase in light intensity (2.5-6.0-10(3) erg-cm(-2)-sec(-1)) . In the presence of DCMU (10(-5)-10(-4) M), hydrogen evolution is not inhibited in the presence of pyruvate or formiate and is inhibited to a less extent in the presence of glucose . According to the results obtained, hydrogen evolution by A . variabilis in the light does not require the action of two photosystems . Inhibition of hydrogen production at significant light intensities is due to the action of oxygen on this process; the rate of oxygen evolution increases with light intensity. Lab Invest, 1976 Nov, 35(5), 430 - 8 Immunologic and cytochemical cell markers in non-Hodgkin's lymphomas; Davey FR et al.; Lymph nodes were obtained from 28 patients with non-Hodgkin's lymphoma and 24 patients without hematologic malignancy . Cases of undifferentiated lymphoma, diffuse histiocytic lymphoma, diffuse and nodular mixed histiocytic-lymphocytic lymphoma, nodular poorly differentiated lymphocytic lymphoma, and diffuse well differentiated lymphocytic lymphoma were analyzed . Touch preparations were stained for nonspecific esterases, peroxidase, Sudan black B activity and with periodic acid-Schiff and Wright-Giemsa reagents . Mononuclear cell suspension from lymph nodes and, in some cases, peripheral blood were tested for spontaneous rosette formation with sheep erythrocytes and for the presence of surface immunoglobulin . The remainder of the lymph node was examined after staining with hematoxylin and eosin . Analysis of the lymphocyte surface markers indicated that 15 cases of various histologic types of lymphoma were B cell proliferations . However, three out of four cases of diffuse poorly differentiated lymphocytic lymphoma and one of seven cases of diffuse histiocytic lymphoma appeared to represent T cell neoplasia . Lymph nodes from four cases of lymphoma representing diverse histologic types were replaced by neoplastic cells devoid of discernible cell markers . In five cases, the distribution of cell surface markers in the malignant lymph node failed to differ from data obtained in the analysis of non-neoplastic lymph nodes . The study indicates that the histopathologic entities recognized in the currently employed classification of lymphoreticular malignancies are heterogeneous . Alterations in the distribution of cell surface markers in the peripheral blood from five of 12 patients indicated involvement prior to demonstrable morphologic evidence of peripheral blood involvement in four patients and bone marrow infiltration in two patients. J Virol, 1976 Nov, 20(2), 384 - 90 Infection of resistant avian cells by subgroup B Rous sarcoma virus; Linial M; Chicken fibroblasts derived from the H & N flock, which have been characterized as resistant to subgroup B avian oncornaviruses in focus assays, can be infected in suspension shortly after trypsinization by subgroup B sarcoma and leukosis viruses . Once cells are plated, resistance to infection reappears rapidly . C/BE cell suspensions obtained by treatment with EDTA instead of trypsin are not as sensitive to infection . Late interference established by preinfection with subgroup B leukosis viruses is not overcome by trypsinization . In addition to C/BE H & N chicken cells, C/ABE RPRL line 7 cells can also be infected by subgroup B viruses shortly after trypsinization; however, none of the cell types can be made sensitive to subgroup E infection . These results are discussed in relation to current information on the genetic control of resistance to avian oncornaviruses. Proc Natl Acad Sci U S A, 1976 Nov, 73(11), 3857 - 61 Subcellular localization of S-adenosyl-L-methionine:tRNA methyltransferases with aminoacyl-tRNA synthetases in human and mouse: normal and leukemic leukocytes; Agris PF et al.; The subcellular distributions of S-adenosyl-L-methionine:tRNA methyltransferases and aminoacyl-tRNA synthetases were investigated with the use of human and mouse normal and leukemic leukocyte cell lines . Differential centrifugation of homogenized cell suspensions produced three pelleted subcellular fractions (nuclear and membrane, microsomal, and postribosomal) and a supernatant fraction . Each fraction was assayed for both methyltransferase activity and synthetase activity . The largest amounts, 40-50%, of total methyltransferase and synthetase activities were localized in either the microsomal or the postribosomal fractions, depending on cell type . In addition, the highest specific activities of these two enzyme systems were found to be present in the microsomal and postribosomal fractions . The psotribosomal fraction from leukemic leukocytes had a methyltransferase specific activity higher than that of the microsomal fraction, while the same two fractions of normal leukocytes had approximately equal activities . Specific activities of aminoacyl-tRNA synthetases were found to be approximately equal for these two fractions, whether they were from normal or leukemic leukocytes . The activity of tRNA methyltransferases and synthetases within the postribosomal fraction of the cytoplasm suggests the existence of high-molecular-weight enzyme complexes for the modification as well as the aminoacylation of tRNA. Arch Dermatol Res, 1976 Oct 27, 256(3), 255 - 60 Long-term tissue culture of epithelial-like cells from human skin (NCTC strain 2544) . I . Measurement of viscosity; Neufahrt A et al.; Viscosities of cell suspensions of human skin epithelium (NCTC strain 2544) were determined by a Wells-Brookfield rotation viscosimeter . A structure viscosity may be postulated by the form of the viscosity curves at different shear-rates . At a cell number of 0.5 X 10(6) cells/ml a viscosity of 1.094 centipoise (25degrees C, shear-rate 230 s-1) could be found . Since a suspension of single cells may be easily formed mechanically (mopping) the NCTC 2544 cells are useful as investigational model regarding the investigation of membrane characteristics. Int J Cancer, 1976 Oct 15, 18(4), 432 - 8 Fc receptor-bearing cells as a reliable marker for quantitation of host lymphoreticular infiltration of progressively growing solid tumors; Kerbel RS et al.; Disaggregated cell suspensions made from transplanted solid tumors, either chemically-induced fibrosarcomas, or spontaneous mammary carcinomas, can contain very high numbers of Fc receptor-bearing cells which are of host origin . Because most types of lymphoreticular cells have Fc receptors, and because T cells--most of which are Fc receptor-negative--appear to infiltrate such tumors only to a very limited degree, the possibility that Fc receptor cells could serve as a reliable and simple marker for host lymphoreticular cell infiltration of solid tumors was tested . This was accomplished by comparing the ratios of Fc rosetting cells to serologically detectable host cells in H2d or H2k haplotype tumor cell suspensions grown in (H2d X H2k)f1 hybrid mice, where host cells could be distinguished from tumor cells by treatment with the appropriate anti-H2 serum . Ratios of 0.8 to 1.08 were obtained for four different tumors including the SaD/2 fibrosarcoma, a CBA spontaneous fibrosarcoma, and the T1699 and CaD/2 mammary carcinomas . Analysis of the results showed that enumeration of Fc rosettes was a reliable host cell maker for at least three of the four tumors tested . The mean non-malignant host cell content of the various tumors, as assessed by anti-H2 cytotoxicity tests, ranged from 23% to 41%. Eur J Immunol, 1976 Oct, 6(10), 757 - 8 The rapid onset of inhibition of antibody-forming cell proliferation upon an increase in density of cellular suspensions cultured in vitro; Gurvich AE et al.; It was shown previously that the development of primary immune response in vitro in the Mishell-Dutton system was strongly inhibited by increased cell densities (Gurvich et al., Immunology 1975 . 28: 271) . In the present work, the rate of onset of this inhibition was studied in C57BL/6 mouse spleen cell suspensions during the phase of exponential increase of antibody-forming cells (AFC) after 3 days' cultivation in vitro with the antigen . It has been shown that the increase in AFC number is very soon inhibited following an increase in the cellular density, the inhibition already being evident within 2 or 4 h . Parallel reduction in incorporation of radioactive precursors in DNA and especially RNA is observed, whereas inhibition of protein synthesis was delayed. Infect Immun, 1976 Oct, 14(4), 872 - 5 Amplified migration inhibition effect; Philp JR et al.; Upon exposure to specific antigen in tissue culture, sensitive lymphocytes released macrophage migration inhibition factor and other lymphokines into the supernatant culture medium . Migration of peritoneal macrophages from nonsensitive animals was inhibited in the presence of such supernatants . However, with previous techniques it was found that an inhibitory effect was present at only low low titers (less than 10(2)) . It is therfore of great interest that by increasing cellular density, the total number of cells being kept constant, inhibitory activity can be amplified by a factor as great as 10(10) . This amplification was observed only when lymphocytes and macrophages were loosely packed, as by spontaneous sedimentation in a conical test tube . The effect was abolished by dispersing the cell suspension in a flat-bottomed flask or, alternatively, by shaking the test tube so that intimate prolonged intercellular contact was prevented. Aust J Exp Biol Med Sci, 1976 Oct, 53(5), 381 - 7 Appearance of cytolytic antibodies in sheep lymph following immunisation with tumour cells . Identification of antibody subclasses; Grant CK et al.; Sheep were immunised with mouse P815 tumour cell suspensions and at intervals afterwards lymph was collected draining from the stimulated lymph nodes . The lymph samples were fractionated by column chromatography into IgM, IgG1 and IgG2 antibody fractions; these were then assayed for cytolytic functions on 51Cr labelled target P815 cells . Complement dependent antibodies were assayed using sheep complement; activity was first detected in the IgM fraction 3-4 days after immunisation and in the IgG1 fraction at 5-6 days . No CDA activity was found in fractions of the IgG2 antibody subclass at any time . Leucocyte dependent antibodies were detected only in the IgG fraction, and they appeared simultaneously in both IgG1 and IgG2 subclasses 5-6 days after immunisation. J Natl Cancer Inst, 1976 Oct, 57(4), 865 - 74 Characteristics of JMV Marek's disease tumor: a nonproductively infected transplantable cell lacking in rescuable Virus; Stephens EA et al.; Cells of the JMV Marek's disease (MD) tumor, originally produced by rapid serial passage of MD lymphoma cells in chickens, were characterized to determine whether they were of host or donor origin and to ascertain certain virus-host cell interrelationships . Differences noted in blood group B surface alloantigens between tumor cells and host lymphocytes indicated a probable nonhost origin (i.e., transplantability) of the tumor . JMV spleen tumors contained predominantly large lymphoblasts bearing MD tumor-associated surface antigen . DNA from JMV tumor cell suspensions hybridized significantly with MD virus cRNA, which indicated that JMV cells contained at least a portion of the MD virus genome . No MD virus was rescued from JMV tumors by techniques suitable for rescue of virus from MD lymphomas . The JMV tumor cells were also devoid of MD virus-specific antigens . These properties differed markedly from those of MD lymphoma cells and make the JMV tumor cell a unique, potentially valuable, tool for further study of oncogenic herpesvirus infection and tumor immunity in the chicken. Biomedicine, 1976 Sep 30, 25(8), 288 - 90 Leucocyte migration fibrinolysis technique (LMFT): description of a method; Coeugniet E et al.; Human leucocytes can migrate in a gel medium consisting of fibrin, 10% horse serum and tissue culture medium 199 . The cells migrate within the fibrin gel mass . The 24 h . areas of migration depend upon the volume and concentration of the cell suspension applied on the fibrin gel . The amount of cells per mm2 migration culture area is less than half the amount used in the leucocyte migration agarose test . The results are reproducible, the standard variation of the migration areas is below 10% . The method is potentially useful for several purposes such as for measuring fibrinolytic and migratory activity of granulocytes, monocytes and lymphocytes and modification of these functions by lymphokines. Int J Cancer, 1976 Sep 15, 18(3), 331 - 8 Inflammatory cells in solid murine neoplasms . II . Cell types found throughout the course of Moloney sarcoma regression or progression; Russell SW et al.; Regressing and progressing Moloney sarcomas, induced in BALB/c mice by the injection of cultured sarcoma cells (MSC)1, were sampled for histologic analysis and then disaggregated using mixtures of trypsin, collagenase and DNAse or collagenase and DNAse alone . The types of inflammatory cells (IC) found in resultant cell suspensions were determined 6, 11, 14 and 18 days post inoculation . Inflammatory infiltrates were composed almost exclusively of three cell types; neutrophils, T lymphocytes and macrophages . The extent to which each was found in tumors was related to the time post inoculation . Neutrophils were part of an early acute inflammatory response seen in both developing regressing and progressing sarcomas . The onset of regression was associated histologically with the appearance within tumors of a mononuclear inflammatory infiltrate . T lymphocytes and macrophages were the principal constituents . A higher percentage of T lymphocytes was recovered at all sampling times from regressing, compared to progressing, sarcomas . During development of the mononuclear inflammatory infiltrate there were relatively more large T cells in regressing, than in progressing tumors, and the percentage of macrophages was higher . Thereafter, the proportion of macrophages in the recovered cell population was approximately the same for both types of tumor . Such equality was more apparent than real, however, since IC were restricted to the peripheries of progressing sarcomas after the acute inflammatory phase, but continued to be found throughout regressing neoplasms . The effective ratio of macrophages and T lymphocytes to tumor cells therefore was much lower in progressing sarcomas than was suggested by percentage figures . The data presented support the concept that T lymphocytes are instrumental in causing the regression of Moloney sarcomas, possibly through interactions with macrophages. Int J Cancer, 1976 Sep 15, 18(3), 322 - 30 Inflammatory cells in solid murine neoplasms . I . Tumor disaggregation and identification of constituent inflammatory cells; Russell SW et al.; Mechanical and enzymatic methods of disaggregating tumors were studied with the goals of (1) minimizing cell losses while (2) maintaining functional and surface membrane markers needed to objectively identify inflammatory cells (IC)1 in resultant suspensions . Application of the principles and methods described makes accurate estimation of the percentage of each IC type present in neoplasms possible for the first time . Compared to purely mechanical means of disaggregating tumors, all enzyme mixtures tested markedly increased yields of viable cells/g neoplasm . Best results were obtained with a combination of collagenase and a protease of broader substrate range (alpha chymotrypsin, papain, pronase or trypsin) . The combination of enzymes that gave the highest yields with the least effect on inflammatory cell markers was trypsin, collagenase and DNAse (TCD) . Because mechanical injury appeared to be the greatest single cause of cell loss (the enzymes themselves had little direct effect), potential sources were identified and either eliminated or minimized . With TCD, depending on the tumor system, cell recovery (measured as DNA recovered in cell suspensions) was as high as 50% and yields were as much as 6.9 X 10(8) viable cells/g tumor . Complete disaggregation was not required to obtain representative IC populations from tumor fragments . Neutrophils, eosinophils and mast cells from disaggregated neoplasms were counted in Giemsa stained cytocentrifuge preparations based on their unique morphologic appearances . Macrophages were identified by their capacity to phagocytose zymosan, a function which proved highly resistant to the effect of enzymes . Flourescent microscopic identification of brain associated thymus antigen (BATA) allowed quantification of T lymphocytes, since this marker was virtually unchanged by enzyme exposure . Surface immunoglobulin (Ig) was stripped from B lymphocytes most rapidly by pronase and chymotrypsin, slowly by trypsin and papain, and not at all by collagenase . Ig positive cells therefore could be quantified in suspensions generated by collagenase or very short (20 min) exposure of fragments to trypsin. Cancer Res, 1976 Sep, 36(9 PT 2), 3471 - 5 Detection of T-cell lymphoma-associated antigens on cord blood lymphocytes and phytohemagglutinin-stimulated blasts; Kaplan J et al.; Absorption studies demonstrate that T-cell lymphoma-associated antigens detected by rabbit antisera to human T-lymphoblast cell lines are present in suspensions of cord blood lymphocytes and phytohemagglutinin-stimulated adult blood lymphocytes in amounts similar to those found in T-cell lymphoma tumor cell suspensions . Smaller amounts of antigen activity are found in suspensions of tonsil cells, thymocytes, and unstimulated adult blood lymphocytes . Little or no antigen activity is found in suspensions of lymphoblasts from patients with other types of leukemia or from B-cell lines . T-cell depletion removes antigen activity from suspensions of normal lymphocytes . These findings suggest that T-cell lymphoma-associated antigens may be fetal antigens expressed by activated T-cells. Allergol Immunopathol (Madr), 1976 Sep-Oct, 4(5), 325 - 32 Localization of T and B lymphocytes in human adenoid, tonsil, appendix and Peyer's patches; Mello JF et al.; T and B lymphocytes were detected in human adenoid, tonsil, appendix and Peyer's patches by adherence to sheep erythrocytes (E) and human erythrocytes sensitized with antibody and complement (HEAC) . The percentages of T lymphocytes in adenoid and tonsil cell suspensions averaged 30.9 +/- 3.4 (S.D.) and 35.8 +/- 6.4 (S.D.) . The percentages of B lymphocytes in the same tissues were 42.5 +/- 11.3 (S.D.) and 40.1 +/- 34 (S.D.) respectively . In adenoid and tonsil tissue sections, B lymphocytes were found in the follicles and T lymphocytes were detected around the follicular areas . The predominant cell population in Peyer's patches and appendix sections was constituted by B lymphocytes. Blut, 1976 Sep, 33(3), 171 - 80 Erythroid colony formation (CFUe) in fetal liver and adult bone marrow and spleen from the mouse; Rich IN et al.; The methyl cellulose modification of the CFUe technique has been applied to 14 day fetal liver and adult bone marrow and spleen from CBA/CA mice . Optimized doses of fetal calf serum, alpha-thioglycerol, erythropoietin and cell suspensions have been obtained from dose response curves in order to standardize the technique . The slopes of the erythropoietin and cell dose response curves indicate a greater sensitivity by fetal liver to the hormone than bone marrow or spleen . The proportion of cells in the DNA synthesis phase of the cell cycle, as measured by the CFUe technique, has been estimated by administering hydroxyurea . Two hours after the drug was injected, 89% of fetal liver cells, 71% of bone marrow cells and 81% of spleen cells were found to be in the S-phase. Immunology, 1976 Sep, 31(3), 443 - 53 Casein-induced experimental amyloidosis . VI . A pathogenic role for b cells in the murine model; Scheinberg MA et al.; The relationship between cellular (B-cell) responses and the development of casein-induced amyloidosis was explored . The main findings are: (1) B-cell abnormalities are more pronounced in amyloid susceptible CBA/J than in amyloid resistant A/J mice; these include increased mitogen responses to SIII, PI:C and DXS, and enhanced primary immune responses to T-independent antigens; (2) CBA/J amyloid spleen cell suspensions appear to be enriched with antibody dependent killer cells and precursors of antibody-forming cells; these abnormalities are not seen in casein treated A/J mice . This hitherto unrecognized proliferation of immunoblasts in casein-treated CBA/J animals raises the possibility that B cell-macrophage interaction may play an important role in the pathogenesis of amyloid disease. Tumori, 1976 Sep-Oct, 62(5), 503 - 15 Electron microscopic observations on T and B lymphocytes from spleens of syngeneic radiation chimaeras; Rebessi S et al.; The ultrastructure of T and B lymphocytes has been examined in long-term syngeneic chimaeras and age-control mice . Spleen cell suspensions from these mice were passed through glass wool columns to obtain pure lymphocyte populations . These cells were then separated into T and B lymphocytes by nylon wool columns, and their purity was tested by cytotoxicity assays with anti-phi serum . Electron microscopic observations on such separated T and B lymphocytes did not reveal morphological differences except when the cells were fully differentiated, either as mature (T2) cells or plasmacells . In particular, T2 cells showed a very high cytoplasmic density, attributable to the presence of a larger number of microfilaments with respect to immature (T1) cells . In long-term chimaeras a significantly larger number of T2 cells was found as compared to age-control mice, and this morphological observation is correlated with the differences in immune reactivity and leukemia incidence previously described in these mice. Can J Microbiol, 1976 Sep, 22(9), 1293 - 9 The respiratory metabolism of Mycobacterium lepraemurium; Kato L et al.; The respiratory metabolism of Mycobacterium lepraemurium isolated from Sprague-Dawley rats lepromata using several substrates was investigated . None of the intermediates of the glycolysis cycle as well as of the tricarboxylic acid cycle except succinate was oxidized by purified whole suspensions of M . lepraemurium . Likewise, many sulfur compounds such as cystine, thiourea, thioacetate, thiodiglycol, mercaptoact and some sulfhydryl compounds, e.g., cysteine, dithioerythritol, dithiorthritol, and penicillamine were readily oxidized by murine bacillary suspensions, whereas thioglycolate, thioglucose, and reduced glutathione were oxidized at a slow rate . Succinate was not or was very poorly oxidized by normal cells probably because of impermeability of the cell wall but the addition of succinate to the cell suspensions frozen for 1 min at -40 degrees C considerably enhanced oxygen uptake over the endogenous value . The oxidation of succinate was unaffected by inhibitors rotenone, atabrine, and amytal but was markedly inhibited by thenoyltrifluoroacetone, antimycin A, 2-N-heptyl-4-hydroxyquinoline-N-oxide, and cyanide . The thiol-binding agents, p-hydroxymercuribenzoate and N-ethylmaleimide were also effective inhibitors of succinate oxidation but the process was not affected by uncouplers dinitrophenol, dibromophenol, pentachlorophenol, and carbonyl-cyanide-m-chlorophenylhydrazone . The results indicated that succinate oxidation by M . lepraemurium was mediated by oxidative enzymes involving an electron transport chain with oxygen as the terminal electron acceptor. J Bacteriol, 1976 Sep, 127(3), 1188 - 96 Energy cost of galactoside transport to Escherichia coli; Purdy DR et al.; Energy reserves of Escherichia coli can be depleted by our previously reported procedure to a level such that even the "downhill" transport of o-nitrophenyl-beta-D-galactopyranoside (ONPG) is completely dependent upon the exogenous energy supply . The ONPG concentration is high externally to the cells and is low intracellular because of the action of cytoplasmic beta-galactosidase . In the present work, depleted cell suspensions have been infused at low, steady rates with glucose and other energy sources while measurements of transport were being made . Comparing the rate of ONPG transport with the rate of introduction of glucose under conditions where the chosen glucose infusion rate limits transport, we find that 89 molecules of ONPG are transported per molecule of fully oxidized glucose . This transport yield is constant over a 6.5-fold range in rate of glucose addition . This constancy over a range of infusion rates implies that transport is the major cellular function under these special conditions . The yield value if 89 is in the agreement with the predicitions of 76 from Mitchell's chemiosmotic theory and constitutes an independent proff of its validity, since all the other proposed mechanisms of engery coupling predict much smaller yields . The lag from the start of glucose infusion into the reaction cuvette, to the extrapolated time at which a steady rate of transport and concomitant hydrolysis are achieved, is short (approximately 1 min) . Similarly, the time after the infusion is stopped until the rate of transport returns to the background rate is also short . The latter implies that the energy metabolism is directed almost entirely to transport and/or other ongoing cellular processes and not to repair or renewal of an energy-independent, facilitated diffusion system. J Endocrinol, 1976 Sep, 70(3), 345 - 59 Purification and characterization of Leydig cells from rat testes; Janszen FH et al.; An LH-responsive Leydig cell preparation (containing 6+/-2% Leydig cells) was obtained by collagenase treatment of rat testis . Centrifugation of this cell preparation through a 13% Ficoll solution for 10 min at 1500 g resulted in a four times purification of the Leydig cells, with a concomitant increases in steroidogenic activity . Addition of 0-2% albumin to the 13% Ficoll solution, adjusted to 280 mosmol/l, resulted in a further twofold purification of the Leydig cells paralleled by a twofold increase in steroidogenic activity . Centrifugation of these Ficoll-albumin-purified Leydig cells through a 6% dextran solution for 2 min at 100 g resulted in a further 1-7 times purification of the Leydig cells . A combination of the two centrifugation steps resulted in a 12-5 times purification of Leydig cells compared with the original crude cell suspension, while an increase in steroidogenic activity of 22-5 times was obtained . This final cell preparation contained 59 +/- 17% Leydig cells (mean +/- S.D., n = 6) . The recovery of Leydig cells was 29% . Collagenase treatment of testes deficient in spermatogenesis resulted in a cell preparation with the same steroidogenic activity as Ficoll-purified cells from normal testes . Centrifugation of these cells through a 13% Ficoll solution gave only a limited increase in the steroidogenic activity . Isopycnic centrifugation of the crude cell preparation on a discontinous Ficoll metrizoate gradient resulted in two discrete peaks of Leydig cells, one peak at a density of 1-039-1-055 g/ml and one at a density of 1-068-1-088 g/ml . Both types of cells produced testosterone . In the presence of LH, cyclic AMP production in both types of Leydig cells increased, but testosterone production was only increased by LH in the "denser" Leydig cells and not in the "light" Leydig cells . No difference in sensitivity to LH could be observed between the Leydig cell preparations of different purity . Using a 60 min pre-incubation period the highest testosterone response was obtained with 100-1000 ng LH/ml . The same maximum testosterone response was obtained with 10-100 ng LH/ml when the pre-incubation period was omitted. Cancer Res, 1976 Sep, 36(9 pt.1), 3171 - 7 Loss or persistence of the differentiated state of simian virus 40-induced hamster tumor cells before and after serial passage in culture; Diamandopoulos GT et al.; The transformed cells that arise from among the hamster epithelial and mesenchymal cells exposed to SV40 in vitro are, as a rule, fibroblastoid and pleomorphic rather than epithelioid . Moreover, the neoplasms that these transformed cells induce in the allogeneic host are spindle cell sarcomas and pleomorphic sarcomas rather than carcinomas . Since this phenomenon may result from cellular dedifferentiation in culture, to the extent that the anaplastic morphology and lack of specialized function can no longer suggest the cell or origin, we investigated the fate of the differentiated state of cells of three types of SV40-induced hamster tumors before and after serial passage in vitro . The tumors evaluated were three reticulum cell sarcomas, three osteogenic sarcomas, and two lymphosarcomas of B-cell origin . Our data demonstrate that reticulum cell sarcoma cells lose their morphological differentiation soon after the original tumors are dissociated into cell suspensions but preserve their phagocytic activity throughout their in vitro passage . Osteogenic sarcoma cells lose their differentiated phenotype and their capacity to form osteoid during but not before their serial passage in culture . Lymphosarcoma cells preserve their lymphoid morphology and their ability to produce immunoglobulin even after many in vitro passages . These results indicate that, in many types of SV40-induced tumors, neoplastic cell dedifferentiation, following serial passage in culture, is responsible to a great extent for the emergence of new cell phenotypes lacking in morphological and functional features characteristic of the cells originally transformed by SV40. Cancer Res, 1976 Sep, 36(9 pt.1), 3113 - 8 Biosynthesis of albumin via a precursor protein in Morris hepatoma 5123tc; Edwards K et al.; The mechanism of albumin biosynthesis was studied in Morris hepatoma 5123tc in vivo and in hepatoma cell suspensions obtained by solubilizing the intercellular matrix with collagenase and hyaluronidase . In the in vivo experiments, L-{-14C}leucine was injected i.v . into rats bearing hepatomas in the muscles of both hind legs . After 14 min, tumors were removed and homogenized . A protein fraction quantitatively precipitable with antialbumin was isolated from the homogenate by acetone fractionation and precipitation with antiserum against serum albumin . This protein fraction was not homogeneous . With the use of 3 consecutive chromatographies on diethylaminoethyl cellulose, a very highly radioactive albumin-like protein could be separated from a large amount of only slightly radioactive albumin . In hepatoma cell suspensions incubated with L-{1-14C}leucine followed by a chase with excess nonradioactive L-leucine, radioactivity was incorporated first into the albumin-like protein and transferred thereafter into albumin, suggesting that albumin was synthesized via the albuminlike protein as precursor . In vivo, 1.8% of newly synthesized hepatoma protein was albumin or its precursor, compared with 1.2% in cell suspensions. Proc Soc Exp Biol Med, 1976 Sep, 152(4), 631 - 4 Role of converting enzyme in the cardiovascular and adrenal cortical responses to (des-Asp1)-angiotensin I; Larner A et al.; (Des-Asp1)-angiotensin I, angiotensin II and III were evaluated for pressor activities in conscious nephrectomized rats and for steroidogenic actions in rat adrenal zona glomerulosa . The pressor effect of this angiotensin nonapeptide was similar to that found with mole-equivalent doses of angiotensin III (one-third as active as angiotensin II) and was significantly attenuated by pretreatment with the 0 . jararaca nonapeptide converting enzyme inhibitor . Hence, (des-Asp1)-angiotensin I is a substrate for converting enzyme in vivo, and the rapid conversion indicates that an alternate pathway for the formation of angiotensin III could exist . (Des-Asp1)-angiotensin I possessed only 0.1% of the activity of angiotensin III as a steroidogenic agent in cell suspensions of rat adrenal zona glomerulosa . Angiotensin I was a weak steroidogenic agent in vitro (1%) and was not blocked by an inhibitor of converting enzyme . Adrenal cells dispersed from the outer zone of the cortex would appear to be devoid of significant converting enzyme activity. Am J Pathol, 1976 Sep, 84(3), 469 - 78 An in vitro model of pancreatic carcinoma . Morphology and in vivo growth; Parsa I et al.; Pancreas rudiments from 13-day rat embryos were cultured in the presence of dimethylnitrosamine (DMN) for up to 10 weeks . Pancreas morphogenesis and differentiation occurred during the first week of culture . Acinar cell degeneration and necrosis began on the fifth day of culture and resulted in almost complete loss of acinar cells, islet cells, and fibroblasts by the end of the third week . This was associated with proliferation of cells without zymogen granules (centroacinar, ductal, or undifferentiated?) . Theses cells formed glandular structures which extended to the surface of the explant . By the end of the fourth week, explants resembled ductal hyperplasia with foci of carcinoma in situ . The distribution pattern of neoplasia in 343 explants examined after 10 weeks of DMN treatment was as follows: 79% resembled ductal cell carcinoma; 9%, ductal hyperplasia; and 3%, acinar cell carcinoma . Nude mice injected with cell suspensions prepared from 10-week-old culture developed subcutaneous nodules . These nodules resembled duct cell carcinoma with desmoplastic reaction. Endocrinology, 1976 Sep, 99(3), 758 - 64 Induction of follicle-stimulating hormone (FSH) receptors in rat ovaries by estrogen priming; Louvet JP et al.; Estrogen pretreatment of hypophysectomized immature rats stimulated granulosa cell proliferation and enhanced FSH binding to the ovary in vivo . The present studies were undertaken to ascertain whether estrogen pretreatment altered the number of specific FSH receptor sites per ovarian cell, resulting in increased FSH binding . Cell suspensions were prepared from the ovaries of groups of rats pretreated with graded doses of 17beta-estradiol . The isolated cells contained specific FSH receptors with high affinity for FSH . The results of these studies clearly showed that the number of specific FSH receptors per ovarian cell was unaffected by pretreatment with graded doses of 17beta-estradiol or with diethylstilbestrol . The enhanced in vivo FSH binding was solely the result of estrogen-induced proliferation of granulosa cells with a fixed number of specific FSH receptors per cell. Ann Immunol (Paris), 1976 Sep-Oct, 127(5), 717 - 31 {The spleen in "nude" mice: an immunological and immunocytochemical study (author's transl)}; Viac J et al.; An immunological and immunocytochemical study to compare spleen cells of Nude (C57B1) and Swiss mice is reported . The percentage of surface immunoglobulins bearing lymphocytes is identical in both strains of animal (40%) . An activated C3 receptor is present in the majority of these cells and has been demonstrated within the germinal centers in both cell suspensions and frozen tissue sections . The population of Fc receptor bearing cells is more heterogenous, but the percentage of these is nearly identical in both Swiss and Nude mice . T-cell specificity is identified with an anti-thymocyte and an antibrain antiserum using an indirect immunoperoxidase method . The percentage of cells detected with the anti-thymocyte antiserum is very low in the spleen of Nude mice (5%) compared to that in the Swiss mice (42%) . In the Nude mice, the anti-brain antiserum detects up to 30% of the spleen cells which may be considered as precursors and, so far, these 30% of cells in the Nude mice are considered as "null" cells, compared to only 20% in normal mice . The localization of the various immunocompetent cells within the spleen tissue is determined by immunofluorescence, immunoenzymology and the EAC rosette test . This study leads to the conclusion that in the periarteriolar areas of the spleen of the Nude mice, immune cells are sparse but that thymo-independent cells are located in the same region in both species of mice. Rev Esp Fisiol, 1976 Sep, 32(3), 187 - 92 Measurement of glycolytic rates in cell suspensions using a recording pH meter; Lopez-Alarcon L et al.; A method for measuring continuously glycolytic rates in cell suspensions, using a recording pH meter, is described . Under the described conditions the method is very exact, sensitive and reproducible . The method can be applied to different cells and different conditions of assay calibrating in each case the pH range, cell concentration range and the ratio of delta protons to delta lactic acid. Eur J Biochem, 1976 Aug 16, 67(2), 527 - 41 Light-induced enzyme synthesis in cell suspension cultures of Petroselinum hortense . Demonstration in a heterologous cell-free system of rapid changes in the rate of phenylalanine ammonia-lyase synthesis; Schroder J et al.; The conditions for protein synthesis in vitro with polyribosomes from cell suspension cultures of parsel (Petroselinum hortense) and a wheat-germ extract were investigated . Two different criteria were used as estimated of the translational activity: (a) the total rate of incorporation of {35S}methionine into acid-insoluble material; (b) the ratio of large (molecular weight greater than 25000) to small (molecular weight less than 25000) peptide products . Depending on which of the criteria was employed, the pH optimum and the optimal concentrations for Tris=acetate, magnesium acetate, KCL, methionine and the wheat-germ extract differed considerably . The translational activity of the polyribosomes (both criteria) was effciently protected by 0.1 M Mg2+ against degradation during the isolation procedure . The rate of synthesis of phenylalanine ammonia-lyase in vitro with the polyribosomes was determined by measuring the incorporation rate of L-{35S}methionine into protein which was precipitable by a rabbit antiserum prepared for the purified enzyme . The immunoprecipitate was analyzed by disc gel electrophoresis in the presence of dodecylsulfate and was shown to contain small amounts of the complete enzyme subunits and relatively large amounts of shorter peptides which were also characteristic for the enzyme . The time course of light-induced changes in the rate of phenylalanine ammonia-lyase synthesis in vitro were investigated during a period of 15 h under two different conditions of induction: the cell cultures were irradiated with ultraviolet light eith (A) continuously or (B) for 2.5 h and then returned to darkness . Although the highest rate of enzyme synthesis was observed somewhat later inexperiment A than in experiment B, the periods of time during which the rate of synthesis increased rapidly were limited in both cases to only a few hours . The results obtained in vitro were identical within the limits of the experimental error with theoretical calculations of the changes in the rate constant of phenylalanine ammonia-lyase synthesis in vivo . These changes were calculated from the corresponding curves for the changes in the enzyme activity under the conditions of induction . The results are in agreement with previous observations suggesting that the induction of phenylalanine ammonia-lyase by light in the parsley cells was a short-term effect whose efficiency was greatly reduced within the 15 h of experimentation, even under continuous irradiation. Int J Cancer, 1976 Aug 15, 18(2), 189 - 96 Interactions of murine leukemia virus (MuLV) with isolated lymphocytes . II . Infections of B and T cells with Friend virus complex indiffusion chambers and in vitro: effect of polyclonal mitogens; Cerny J et al.; The infection of isolated B and T cells by a murine leukemia virus (Friend) MuLV-F) was studied both in vitro and in vivo with an implanted diffusion chamber system . Lymphocytes were obtained from pools of normal spleen cells by filtration of the cell suspension through a nylon-wool column . The purity for both Ig positive and theta-positive cells varied between 85% and 90% in the B-cell and T-cell fractions; both lymphocyte fractions responded very well to stimulation with their respective specific polyclonal mitogens, bacterial lipopolysaccharide (LPS) and concanavalin A (Con A) . Lymphocytes were infected by incubating pelleted cells in 2-6 x 10(4) FFU MuLV for 1 h at 4 degrees C and were then cultured for 5-10 days . Cells releasing infectious MuLV were enumerated as infectious centers (IC) . IC were really detectable in the cultures of infected B-cells but none were found in the T-cell cultures . Addition of LPS to the culture medium increased the number of IC in B-cell fractions up to 1,000-fold . Furthermore, in T-cell cultures with LPS, IC also appeared in number which approximately correlated with the contaminating Ig+ cells of the T-cell fraction . In contrast, Con A had no consistent effect on the infection of either B or T cells . In the absence of MuLV-F, mitogenic stimulation alone did not elicit any endogenous IC . In subsequent experiments, purified lymphocytes were infected in diffusion chambers in vivo . The number of IC in infected B cells increased 1,000-fold as compared to infection in tissue culture . The peak of infection at 10 days was followed by a slight decline . Infected cells were also found in diffusion chambers containing T-cell fractions; these IC had very similar kinetics to those in B-cell-containing chambers, but their number was 10 times lower, suggesting that the infected cells were B cells, which comprised about 10% of the T-cell fraction . The virus-related antigens were detectable by immunofluorescence on the membrane of cells recovered from B-cell-bearing chambers but not on cells from T-cell-bearing chambers. Int J Cancer, 1976 Aug 15, 18(2), 197 - 204 Interactions of murine leukemia virus (MuLV) with isolated lymphocytes . III . Alterations of splenic B and T cells in Friend virus-infected mice; Cerny J et al.; Lymphoid tissues of mice infected with murine leukemia virus (Friend) (MuLV-F) were examined for the presence of cellular markers of MuLV-F infection . The Friend virus-associated cell membrane antigen (FVMA) and the virus group-specific antigen (GSA) were detectable on cells from the spleen and, to a lesser degree, on cells from the bone-marrow . In contrast, neither FVMA nor GSA was found in cells from the thymus . Alterations in the B-cell and T-cell spleen populations of MuLV-F-infected mice were then studied . The proportion of Ig-positive cells declined from the initial 45% (in non-infected controls) to about 10% after 2 weeks of infection . A similar decline of theta-positive cells was noted . However, complement-bearing cells (EAC rosettes) declined even more rapidly and became undetectable in the second week after infection . The treatment of spleen cells from MuLV-F-infected mice with anti-FVMA serum plus complement in vitro reduced the number of detectable Ig-positive cells, specifically, whereas the number of theta-positive cells remained unchanged . Furthermore, B and T cells from spleens of infected mice were separated on an affinity column with anti-Ig antibody-coated beads . The initial cell suspension contained about 45% FVMA-positive cells, about 40% Ig-positive cells and about 40% theta-positive cells . Ig+ cells were retained on the column . The theta-positive cell fraction was collected in the eluate and contained very few FVMA-positive cells with some "null" cells . Most of the FVMA-positive cells were retained on the column, which strongly suggested that they were B cells . These results confirm the previous experiments which showed the selective infections of purified splenic B cells by MuLV-F in cultures. Biochim Biophys Acta, 1976 Aug 13, 440(2), 429 - 47 Membranes of Rhodopseudomonas sphaeroides . IV . Assembly of chromatophores in low-aeration cell suspensions; Niederman RA et al.; Chromatophore membrane formation was induced in low-aeration suspensions of Rhodopseudomonas sphaeroides and highly purified chromatophore preparations were isolated at various intervals between 4 and 18 h . The levels of several functional components associated with the isolated strucures were investigated . B-875, the light-harvesting bacteriochlorophyll complex associated with the reaction center, was preferentially inserted into the chromatophore membrane during the early stages of induction, and thereafter its levels reached a steady state; b- and c-type cytochromes were also maintained at essentially constant levels . In contrast, the levels of B-850, the accessory light-harvesting bacteriochlorophyll, together with its associated protein, continued to increase throughout the induction process . Increases in the levels of the major carotenoid component followed a similar course . These findings are consistent with a stepwise assembly mechanism for associated bacteriochlorophyll and protein components and suggest that separate regulatory mechanisms control the levels of functionally essential and accessory components within the membrane. Clin Exp Immunol, 1976 Aug, 25(2), 319 - 27 Rosette-formation with mouse erythrocytes . II . A marker for human B and non-T lymphocytes; Gupta S et al.; Peripheral blood lymphocytes from healthy control subjects were studied for spontaneous rosette-formation with mouse erythrocytes . The mean percentage of mouse erythrocyte rosette-forming cells (MRFC) in sixty-three subjects was 7.4X3.5; after neuraminidase treatment olymphocytes the mean percentage of mouse erythrocyte rosette-forming cells(NMRFC) was 17.2X5.6 . Thymus cell suspensions made only occasional rosettes with mouse erythrocytes . There was no effect of neuraminidase treatment of thymocytes on mouse erythrocyte rosette-formation . Double labelling experiments with other cell surface markers confirmed this binding of mouse erythrocytes to be a B-cell characteristic . Monocytes and granulocytes do not bind mouse erythrocytes . It appears that there is a distinct receptor for the binding of mouse erythrocytes on the surface of lymphocytes carrying surface immunoglobulins; neuraminidase treatment of lymphocytes permits this binding to occur also on virtually all the non-T cells B cells plus the third population cells). J Natl Cancer Inst, 1976 Aug, 57(2), 345 - 8 Role of donor immunocompetent cells in allograft rejection; Jacobs BB et al.; Allotransplantable lines of the BALB/c (H-2d) Leydig cell tumor C4092 were established in DBA/1 (H-2q) mice after maintenance in organ culture . These modified tumors had reduced immunogenicity but were recognized and rejected by previously sensitized DBA/1 mice . The addition of 3.4 X 10(3)-1 X 10(5) peritoneal exudate cells (PEC) from untreated BALB/c mice to cell suspensions of the modified tumor did not restore Immunogenicity when the cell mixture was Inoculated sc Into DBA/1 recipients . However, when equivalent numbers of PEC or spleen cells were inoculated iv into animals receiving the tumor cell suspension sc, the acceptance of the tumors was significantly reduced or prevented . This indicated that the BALB/c lymphoid cells inoculated iv stimulated the immune system of the host, which in turn recognized the tumor cell inoculum . The lack of effect of the BALB/c lymphoid cells when admixed with the tumor did not support the assumption that the loss of passenger lymphoid cells was responsible for the allotransplantability of grafts after organ culture explantation . Additional evidence was presented that supports our view that the reduced immunogenicity was associated with a phenotypic modification of the tumor cells per se. Tsitologiia, 1976 Aug, 18(8), 1003 - 7 {Effect of uncoupling agents and benz (a) pyrene metabolites on respiration in L cells and normal mouse embryonal fibroblasts}; Riabykh TP et al.; A method is proposed for a polarografic study of the respiration of cell suspensions obtained from monolayer cultures of L-cells and from normal embryonic fibroblasts of mice (C3HA line) . 6-hydroxybenzo(a)pyrene (6-OHBP) . The metabolite of the carcinogenic hydrocarbon benzo(a)pyrene, was shown to be a strong uncoupler of of oxidation and phosphorylation of cell suspensions . Its effect was influenced by the presence of calf serum in the incubation media . Possible relationships between the toxic effect of 6-OHBP on monolayer cultures of normal and tumor cells, and its effect on cell energetics are discussed. Beitr Pathol, 1976 Aug, 158(3), 255 - 86 {Sarcoma 180: growth and regression . Comparative investigations using flow-through and scanning cytophotometers as well as histological, cytological and autoradiographic techniques (author's transl)}; Stecher G et al.; Material and methods: The growth and the regression of the experimental tumor S 180 was investiaged by volume measurements and by cytophotometric studies on 50 mice in each of a pilot and in the present main experiment . In addition, histological, cytological and cytokinetic examinations were performed . Results and discussion: Beginning on day 18, a spontaneous regression of the tumor was found in 20% and 38% of the animals, respectively . DNA-measurements were performed with a scanning-microspectrophotometer on tumor tissue imprints and tumor cell suspensions and were also carried out with a flow-through cytophotometer ICP on suspensions . DNA-histograms were plotted on days 4, 7, 12, 18, 20, 22, 26 and 29 after the transplantation . These revealed a constant position of the maxima at 2C for normal cells and at 4C and 8C for the tumor cells . In particular, by measuring a large total number of 4,968,000 nuclei with the ICP, distinct changes were found in the proportion of the single nuclei classes during growth and spontaneous regression . This technique also enables a quantitative measurement to be made of the DNA of cell debris from necrosis . With growing tumos, the proportion of the tumor cells increased to a maximum of 65% on day 18 and decreased to 37% on day 29 . The DNA of cell debris grew from 8% to 47% . The proportion of normal cells was only 10% to 15% in the final phases . In tumors with a spontaneous regression the proportion of the tumor cell nuclei was 20% on day 18 and 11% on day 29 . The proportion of the DNA of cell debris was again about 45% . The proportion of normal nuclei was greatly increased to 45% . Histological and cytological evidence together with measurements of the areas of nuclei and incorporation of 3H-TdR confirmed that the normal cells, the number of which increased during spontaneous regression were cells of a fibrovascular granulation tissue . While the rate of the DNA-synthesis of growing tumors continously decreased with age there was also a rapid decrease of the labelling index of spontaneously regressing tumors between days 12 and 18 . In the final phase only the nuclei of the granulation tissue were labelled. Transplantation, 1976 Aug, 22(2), 101 - 7 Influence of separation techniques on the distribution and function of lymphocyte subpopulations . A comparison of three techniques; Mookerjee BK; The effect, if any, of three different lymphocyte separation techniques on the composition and functional characteristics of the purified cell suspensions has been studied . Each separation technique was shown to yield a cell population with highly specific and reproducible characteristics . The Ficoll-Hypaque techniq