Culture medium

In Vitro, 1976 Feb, 12(2), 115 - 9
Primary culture and maintenance of steroid-secreting human placenatal monolayers; Roy PW et al.; A simple method is described for primary culture and for maintenance of hormone-producing cells from normal human placenta . A consistent yield of cells was obtained and an average survival of 3 to 4 months in culture using 1 mm3 explants from the most vascular area of the placentas . These explants were placed in a variety of culture media in 30 ml flasks and incubated at 37 degrees C in an atmosphere of 5% CO2 and 95% air . The best yields in terms of cell growth were observed with Eagle's MEM (minimum essential medium) with supplements of horse serum and fetal calf serum or human cord serum . (Ham's F-10 with supplement of horse serum and fetal calf serum supports growth for the longest period and media containing human cord serum had the best yield of steroids.

Biochemistry, 1976 Feb 10, 15(3), 521 - 8
Regulation of 3-hydroxy-3-methylglutaryl coenzyme A reductase activity and the esterification of cholesterol in human long term lymphoid cell lines; Kayden HJ et al.; The regulation of the rate-controlling enzyme in cholesterol biosynthesis and of the incorporation of {14C}oleate into cholesterol esters were studied in established lymphoid cell lines from normal subjects and compared with that of eight patients with genetic abnormalities of lipid metabolism . The activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase, the rate-controlling enzyme in cholesterol biosynthesis, increases in lymphoid cell lines derived from normal subjects after the culture medium is changed to a lipid deficient medium and reaches peak activity after 48 hr . The addition of whole serum and of low density lipoproteins to cell lines derived from normal subjects suppressed 3-hydroxy-3-methylglutaryl coenzyme A reductase activity by 50%, but failed (almost completely) to suppress the activity in the lymphoid cell lines derived from two patients with homozygous familial hypercholesterolemia . When 7-ketocholesterol was added, the activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase was markedly suppressed in both normal and abnormal lymphoid cell lines . Lymphoid cell lines derived from patients presumably heterozygous for familial hypercholesterolemia were difficult to distinguish from normal cells in these studies . The incorporation of {14C}oleate into the fatty acid fraction of cholesteryl esters was stimulated by the addition of the low density lipoproteins to the culture media of the lymphoid cell lines derived from the normal human subjects . The lymphoid cell lines derived from the patients with homozygous familial hypercholesterolemia showed no increase in {14C}oleate incorporation into cholesteryl esters even when a fourfold amount of low density lipoprotein was added to the media; a modest increase in {14C}oleate incorporation was observed in lymphoid cell lines from patients with heterozygous familial hypercholesterolemia . The results of these studies in lymphocyte cell lines are compared with the findings in cultured human fibroblasts obtained from normal subjects and from patients with homozygous familial hypercholesterolemia . Studies of the regulation of cholesterol biosynthesis in the apparently permanent lymphoid cell line maintained in suspension culture offer certain advantages over cultured skin fibroblasts, and, in addition, provide a second tissue for the study of genetic abnormalities from the same patient.

Endocrinol Jpn, 1976 Feb, 23(1), 65 - 73
In vitro biosynthesis of human chorionic follicle stimulating hormone; Maruo T; In order to explore the possibility that human chorionic FSH (hCFSH) may be synthesized in vitro by the placenta and secreted into the culture media, chorionic tissue of the first trimester was cultivated in the radioactive medium prepared byadding 3H-proline and/or 14C-glutamic acid . Purification of biosynthesized hCFSH from the media was carried out by a combination of Sephadex G-100 gel filtration, DEAE-cellulose chromatography and polyacrylamide disc-gel electrophoresis...

J Clin Microbiol, 1976 Feb, 3(2), 212 - 3
Comparison of sodium amylosulfate and sodium polyanetholsulfonate in blood culture media; Hall MM et al.; A comparison between sodium polyanetholsulfonate and sodium amylosulfate in unvented vacuum blood culture bottles containing tryptic soy broth was made with 5,800 sets of blood cultures . No statistically significant differences in isolation rates of bacteria were noted.

Gann, 1976 Feb, 67(1), 131 - 5
Alpha-fetoprotein and albumin produced by subclonal cell population of the ascites hepatoma AH-66 in vitro; Isaka H et al.; Subclonal cell populations were prepared from a clonal line of the rat ascites hepatoma AH-66 in vitro, using the agar plate culture method . alpha-Fetoprotein and albumin concentrations in culture media of these subclones were determined by 125I-radioimmunoassay and single radial immunodiffusion method, respectively . Results demonstrated that the AH-66 clone was a complex of cells with varying producibility of alpha-fetoprotein and albumin . All the subclones showed varied and distinct production of alpha-fetoprotein, but not all the subclones produced detectable levels of albumin.

J Clin Microbiol, 1976 Jan, 3(1), 47 - 8
Recovery of pathogenic fungi from clinical specimens submitted for mycobacteriological culture; Roberts GD et al.; A total of 332 major pathogenic fungi were isolated from specimens cultured onto both fungal culture media and media used for culturing mycobacteria from January 1968 to June 1975 . Only 72 (21.7%) fungi were recovered on media used for culturing mycobacteria . The effect of sodium hydroxide treatment was evaluated and shown to be detrimental to the recovery of fungi . It is recommended that clinical laboratories not rely on mycobacteriological cultures to recover fungal pathogens.

Clin Endocrinol (Oxf), 1976, 5 Suppl, 387S - 396S
The ectopic secretion of calcitonin by lung and breast carcinomas; Coombes RC et al.; Many patients with advanced non-thyroid malignancies have elevated plasma immunoreactive calcitonin concentrations . Breast and bronchial carcinomas contain immunoreactive calcitonin and an epidermoid bronchial carcinoma has been shown to produce immunoreactive calcitonin in vitro . We have established monolayer cultures of breast carcinomas and eight out of fifteen consecutive carcinomas released immunoreactive calcitonin; some released HCG (human chorionic gonadotrophin) or CEA (carcinoembryonic antigen) . In addition, a primary human breast carcinoma has been shown to release and contain calcitonin after being passaged in 'nude' mice over 1 year . Chromatography of extracts and culture media of a bronchical carcinoma demonstrated that, in contrast with the other tumours, it secreted a form or forms of calcitonin having size, charge and immunological differences when compared to calcitonin M . Preliminary evaluation of plasma immunoreactive calcitonin estimations in patients with breast carcinoma showed that twenty-three out of twenty-eight patients with metastatic disease had elevated plasma calcitonin concentrations, whereas only one out of thirteen with localized disease had high levels.

Cytobios, 1976, 17(66), 87 - 102
On formation of melanosomes in a cultured melanoma line; Schjeide OA et al.; Melanosomes are present in abundant numbers in HPM-73 melanoma cells maintained in surum-containing cultures . However, withholding of serum for 3 days caused severe retardation of development of these organelles . Addition of chondroitin-4-sulphate (0.4 mg/ml) to serum-containing culture media for 3 days resulted in greater numbers of premelanosomes and melanosomes relative to mitochondria . The same amount of chrondroitin-4-sulphate (0.4 mg/ml) added to serum-free cultures also resulted in an increased ratio of premelanosomes and melanosomes to mitochondria but, within the 3-day period many of these organelles did not attain the pattern of maturity evident in melanoma cells reared in serum-supplemented media . By exploitation of slowed rates of development of melanosomes in cultures (especially, but not exclusively, in serum-free samples) it was possible to examine various early developmental forms of these organelles . It was found that in this cell line mitochondria appear to be involved in the process of melanosome formation.

IARC Sci Publ, 1976, (13), 253 - 60
{Capacity of cells in culture to a accumulate polycyclic aromatic hydrocarbons at microspectrofluoremetric detectable levels . Preliminary results}; Salmon JM et al.; In the course of more general microspectrofluorometric studies on intracellular metabolic control mechanisms, the behaviour of living cells was investigated in culture media containing traces of Benzo(a)Pryene (BP) difficultly recordable by spectrofluorimetry . A rapid transfer of BP to the intracellular phase with accumulation was observed . In this manner the recording of the hydrocarbon fluorescence emission spectrum from one single cell became very easy . This fact has led to the investigation of the experimental conditions required for a quantitative detection of trace amounts of aromatic hydrocarbons, and the results presented here are the initial finding of such analysis . In a first part, the requirement for a recording of the complete fluorescence emission spectrum will be emphasized as a condition for a quantitative results . The second part of the presentation is devoted to observations effectively made on fluoresence emission spectra recorded from cells grown in the presence of traces of BP . Due to the aim pursued it is evident that the cells selected for this type of studies should exhibit a relatively slow metabolization of the hydrocarbon . Two significant facts emerge: (a) in most cases the fluorescence spectrum recorded from BP-medium grown cells is not identical to that of BP in solution (less defined structure, displaced maxima, and modified relative intensities of maxima) . The spectrum observed in the BP medium grown cells is apparently the result of two spectra, one corresponding to a free BP fraction, the other to a BP fraction interacting with cellular constitutents . (b) When the cells were maintained for periods up to one to three months in presence of low amounts of BP, it was noticed that the BP fractions "interacting" seemed relatively more significant than in cells non "adapted"to BP . Both results are evidently subject to biological intrpretations outside the scope of this communication . However, they permit to draw the attention on the fact, that the quantitative determination of the hydrocarbon in the living cell can be achieved only after definition of a sufficiently precise experimental protocol.

Arch Environ Contam Toxicol, 1976, 4(4), 483 - 91
Absorption and metabolism of a selective insecticide, 2,2-dimethyl-2,3-dihydrobenzofuranyl-7 N-dimethoxyphosphinothioyl-N-methylcarbamate, in bean plants; Krieger RI et al.; The absorption and metabolism of 2,2-dimethyl-2,3-dihydrobenzofuranyl-7 N-dimethoxyphosphinothioyl-N-methylcarbamate (PSC), a selective, insecticidally active carbofuran derivative, was studied in red kidney bean plants . PSC was absorbed from hydroponic culture media and was translocated throughout the plants . PSC was metabolized to carbofuran, 3-hydroxycarbofuran, and 3-ketocarbofuran and these materials accounted for 25% of the recovered radioactivity . Each of these compounds are more toxic to mice than the parent compound . Additionally at least four other oxidized, hydroxylated and/or hydrolyzed metabolites were formed.

Tex Rep Biol Med, 1976, 34(1), 45 - 50
On the nature of the defect in cystic fibrosis; Conover JH et al.; Sera and lymphocyte culture media derived from cystic fibrosis (CF)-affected and carrier subjects contain ciliary dyskinesia factor (CDF) detected by our rabbit tracheal bioassay . In addition, we also find CDF in fibroblast media from these same donors and in amniotic fluid cell media derived from CF carrier or affected fetuses . In these latter instances, the media were inactive in the bioassay, but became active when mixed with purified IgG . In all instances, CDF activity was eliminated by the addition of anti-IgG . We have separated a low molecular weight fraction, between 1,000 and 10,000 M.W., from CF sera and culture media which is inactive in the bioassay until IgG is added . Presumptive and indirect evidence indicates that this fraction behaves similarly to the complement derived anaphylatoxin C3a . In addition, we have found activity in sera from CF patients and, to a lesser extent, carriers that induces degranulation of cytochalasin-B-treated human polymorphonuclear leukocytes . Since this activity appears to be in a different molecular species from that containing CDF, we postulate that the primary defect in CF is the deficiency of an enzyme whose substrates include a family of membrane-active molecules.

J Immunol Methods, 1976, 12(1-2), 91 - 102
Short-term microcultures of lymphocytes from Chinese hamster peripheral blood; de Jong B et al.; A microtechnique for the culture of Chinese Hamster lymphocytes is described using Cooke microtiter plates with 100,000 leucocytes in a culture volume of 0.1 ml and a culture time of three days . The culture media used were RPMI 1640 and Trowell T8 supplemented with 20% foetal calf serum (FCS) and 2 mu PHA . The cells were harvested with a Skatron cell culture harvester using glass fibre filters . Various technical aspects of the lymphocyte cultures from the Chinese Hamster are described . The relevance of changes in culture conditions to the variability of culture results was analysed for PHA and FCS concentrations, different culture media, cell concentration, vessel shape and culture duration.

Z Allg Mikrobiol, 1976, 16(6), 425 - 35
Metal accumulation by bacteria with particular reference to dissimilatory sulphate-reducing bacteria; Jones HE et al.; Dissimilatory sulphate-reducing bacteria, genera Desulfovibrio and Desulfotomaculum, exhibit a superior ability, over assimilatory organisms, to extract three amounts of metals from culture media . This property does not appear to be solely a function of the presence of H2S . In media containing elevated amounts of Fe, electron dense particles, provisionally identified as FeS, are deposited within the cells of dissimilatory bacteria.

Dev Biol Stand, 1976, 33, 89 - 92
Efficacy of a live oral typhoid vaccine in human volunteers; Hornick RB et al.; A live oral attenuated vaccine, lacking the enzyme epimerase, has been given with complete safety to 173 men . Two preparations of the vaccine, one with and one without galactose in the initial culture media, led to somewhat differing results . Vaccine A, prepared with galactose, was more readily identified in the stool specimens of its recipients and was more likely to provoke an O antibody response than was vaccine B . Furthermore, the clinical protection from typhoid fever was significant in the vaccine A group, whereas not so with the vaccine B counterpart . The protection afforded by vaccine A exceeds that of any of our previous candidate strains . It is perhaps not presumptuous to imagine that such a vaccine would be equally effective in an area endemic for typhoid fever, where the vaccine might act as a booster effect in a previously exposed population . Remaining questions to be answered include the duration of the protection, and the efficacy of a lyophilized preparation.

Pathol Biol (Paris), 1976 Jan, 24(1), 61 - 6
{Iso-hormones : molecular forms of human chorionic somatomammotropin (HCS) (author's transl)}; Belleville F et al.; The chorionic somatomammotropin hormone extracted from the human placenta exists in several molecular forms: Analytical electrophoresis on polyacrylamide gel permits separation of a highly anodic migration form : form 1 and another form migrating slightly faster than albumin : form 2 . These two forms are active as measured by radioactive immunological analysis, form 2 being about 25 times more active than form 1 . The two forms are mutually interconvertible . The two forms may also be separated by filtration on Sephadex G-50 . However, they do not differ in molecular weight, they have the same coefficient of apparent diffusion, measured by analytic ultracentrifugation . Glutaraldehyde and 8 M urea do not modify their electrophoretic or chromatographic behaviour . On the other hand, the two forms differ in their tertiary structure, with the modification depending on the greater or lesser degree of oxidation of the intra-chain disulfide groups . The two forms also exist in placental culture media and the incorporation of tritiated leucine occurs preferably in form 1, The physiological significance of the two hormone pools is not clarified.

Tex Rep Biol Med, 1976, 34(1), 73 - 82
Immunochemical studies of the plasma and cultured fibroblast media fractions containing the cystic fibrosis ciliary inhibitor; Harper BL et al.; The cystic fibrosis ciliary inhibitor (CFCI) has been fractionated from plasma of cystic fibrosis (CF) homozygotes and from the media of cultured fibroblasts derived from CF homozygotes . Plasma and fibroblast media from normal controls have been fractionated in an identical manner . Fractions from plasma and fibroblast culture media that demonstrate ciliary inhibitory activity contain several proteins in a molecular weight range of approximately 5,000-11,000 . These proteins have been partially characterized by immunochemical analysis with antisera to 33 human serum proteins . Immunological determinants of albumin, C3 (but not C3a), C4, C5, alpha1-lipoprotein, beta-lipoprotein, beta2-microglobulin and immunoglobulin light chains have been detected by hemagglutination in fractions of CF plasma that inhibited ciliary activity and in analogous fractions from normal sera . None of the proteins were detected in media of cultured fibroblasts from either genotype . Since the same proteins and protein fragments were identified in both CF and normal plasma fractions, and were not detected in CF fibroblast media, it appears that none of these proteins can be identified as the CFCI . Identification of these proteins will permit further purification of the CFCI by immunochemical methods.

Birth Defects Orig Artic Ser, 1976, 12(3), 305 - 12
Phenotypic expression of galactokinase deficiency in heterozygous and homozygous subjects: in vivo and in vitro studies; Benson PF et al.; Cultured fibroblasts derived from a patient homozygous for galactokinase deficiency, his parents, and controls had similar rates of growth in culture media where the only hexose was glucose . However, in media where the only hexose was galactose there was almost no growth of homozygous mutant cells or of maternal heterozygous cells and slight growth of paternal heterozygous cells . Growth of control cells was initially slow, but after a lag period (which coincided with increasing galactokinase activity) growth reached approximately the same levels as in glucose medium . In all cell lines there was a direct relation between the degree of enhancement of galactokinase activity and the ability of cells to adapt to growth in media where the only hexose was galactose . Erythrocyte galactokinase activities in a series of 24 children children with congenital cataracts aged 2-16 years were similar to those in 26 controls . One child in each of the cataract and control groups had 40-50% of mean control activity and was considered to be a potential heterozygote . Galactokinase deficiency (homozygous and heterozygous) is considered to be an uncommon cause of childhood cataracts . Nevertheless, it is an important cause since early dietary treatment can prevent or reverse lens opacities . The heterozygous state may be expressed phenotypically in the patient by the appearance of cataracts and in cultured cells by their defective growth in media where galactose is the only hexose.

Rev Ig Bacteriol Virusol Parazitol Epidemiol Pneumoftiziol Bacteriol Virusol Parazitol Epidemiol, 1976 Jan-Mar, 21(1), 49 - 53
{Use of culture media with sheep serum for maintenance and isolation of Leptospira strains}; Nicolescu M et al.; A study was carried out on the effects of a culture medium prepared with sheep serum inactivated at 68 degrees C on Leptospira cultures . Good results were obtained showing that the medium with sheep serum can be used for the cultivation of Leptospira in view of the preparation of antigens for microscopic agglutination and complement fixation . In experiments on the isolation of germs from organocultures and haemocultures the proportion of positive results obtained with sheep serum was smaller than with the media containing rabbit serum.

Exp Pathol (Jena), 1976, 12(3-4), 149 - 58
Acid mucopolysaccharides in fibroblast cultures . 1 . Influence of cell density, pH-value and lactate concentration on the MPS distribution pattern; Kittlick PD et al.; From cells and culture media of embryonic rat fibroblasts (1st subculture) the acid mucopolysaccharides were isolated and fractionated . The per cent calculation of the 6 fractions was based on the content of glucuronic acid . The cultures were maintained as follows: 0.5 X 10(6) to 3 X 10(6) cells were examined in Demeter flasks at pH 7.4 or 6.6; lactate concentration was enhanced to 100 mg% . The amount of each fraction was correlated with the cell density (linear regression) . The pH-value and lactate concentration in connection with cell density proved to be important factors in the modification of the MPS distribution pattern.

Diabete Metab, 1975 Dec, 1(4), 227 - 34
Biosynthesis of basement membrane collagen in cultures of renal glomerular and tubular epithelial cells; Foidart-Willems J et al.; Confluent cultures of renal glomerular or tubular epithelial cells were incubated with {14C} proline and {3H} lysine . The incorporation rate of both radioactive precursors was found to be linear for up to 12 h . The synthesis and secretion of basement membrane collagenous polypeptides was demonstrated by the presence in the culture media of non-dialyzable 4-hydroxy {14C} proline and hydroxy {3H} lysine . After gel filtration of the culture media on Sephadex G-100 columns, glomerular and tubular basement membrane polypeptides were identified in the chromatographic fractions by radioimmunoassay . They were further purified by affinity chromatography, using Sepharose cyanogen-bromide coupled with specific rabbit anti-human glomerular or tubular basement membrane antibodies . Absorbed labelled membrane polypeptides were eluted from the Sepharose by acidic medium at 4 degrees C . This membrane material represented 3-4% of the total proteins synthesized by glomerular and tubular cells . The glomerular and tubular basement membrane polypeptides purified by affinity chromatography exhibited a molecular weight of approximately 140,000; 80% of the total hydroxy {3H} lysine was recovered as glucosyl-galactosyl-hydroxy {3H} lysine . Analysis of the carbohydrate content of labelled basement membrane polypeptide chains originating from glomerular or tubular cells incubated with {14C} glucose of {14C glucosamine indicated the presence of glucose, galactose, mannose, glucosamine, and galactosamine . No fucose, mannosamine or sialic acid were detectable . The data demonstrate that glomerular and tubular epithelial cells are able to synthesize basement membrane collagenous polypeptides in culture . This property might provide a useful tool for the study of the biosynthesis of similar material by diseased kidneys.

J Gen Microbiol, 1975 Dec, 91(2), 233 - 40
The inhibitory action of fatty acids on the growth of Escherichia coli; Fay JP et al.; The effect of fatty acids on Escherichia coli K12 was dependent on the source of the inoculum, the growth phase and the washing of the bacteria . The effects of saturated fatty acids from C4 to C16 and oleic acid at two concentrations (0-I and 0-4%, w/v) were determined on E . coli K12/154 growing exponentially in five different culture media . Depending on the media, 0-I % fatty acids increased the doubling times of the cultures by up to 96 % . Fatty acids of medium chain length (C6 to CII) at 0-4 % produced a decrease in cell concentration, nonanoic and decanoic acids being the most effective . A correlation was found between the decrease in cell concentration and the loss of viability of the culutres after addition of 0-4 % decanoic acid, with stationary-phase bacteria being affected more than those from exponential-phase cultures . Experiments carried out with E . coli B and C gave results similar to those obtained with E . coli K12/154.

Infect Immun, 1975 Dec, 12(6), 1242 - 51
Extracellular iron acquisition by mycobacteria: role of the exochelins and evidence against the participation of mycobactin; Macham LP et al.; Mycobacterium bovis var . BCG was grown under iron-deficient conditions in the presence and absence of 1% Tween 80 . Mycobactin, the iron iron ionophore of mycobacteria, was found solely within the bacteria grown in the absence of Tween, but low concentrations (0.75 mug/ml) of it appeared in the medium in the presence of the surfactant . Both types of medium contain agents, named exochelins, which could solubilize iron . 55Fe added to spent culture media was recovered only chelated to these compounds . Two exochelins were detected, isolated, and purified . Neither were precursors or breakdown products of mycobactin . In the desferri-form, exochelin MB-2, the major component, reversed the inhibitory effect of serum on the growth of BCG, and in their ferri-forms exochelins MB-1, MB-2, and MS (from Mycobacterium smegmatis) stimulated the growth of their producing organism in the presence of serum . Exochelin MB-2 could physically remove iron from ferritin, and BCG used ferritin as a source of iron during growth even when ferritin was separated from the bacteria by a dialysis membrane . As solutions of the exochelins were freely dialyzable, whereas solutions of mycobactin, even in Tween, were not, only exochelin could have been active in this experiment . The exochelins are proposed as the functional extracellular iron-binding agents of BCG and other mycobacteria, the role of mycobactin being confined to that of a cell wall iron transporter.

Diabetologia, 1975 Dec, 11(6), 527 - 34
Human islet cell adenoma: metabolic analysis of the patient and of tumor cells in monolayer culture; Adcock K et al.; Cell cultures were established from a benign pancreatic islet adenoma . Over 200 muU/culture/day immunoreactive insulin were found in culture media . Cultures with medium 199 released insulin for about 2 months; those with medium F12K were maintained for over 7 months, and have been successfully subcultured . Increasing culture medium glucose to 326 mg per 100 ml, alone or with leucine (10 mM) or theophylline (2 mM), failed to increase insulin release above baseline . Studies in the patient prior to surgery using oral glucose, leucine, beef meal, intravenous tolbutamide, and glucagon failed to increase plasma insulin and thus were consistent with cell culture responses . Extracts of tumor tissue contained 23% proinsulin-like material; high insulin containing samples of culture medium had 5% proinsulin and less than 40 pg glucagon/ml . Aldehyde fuchsin positive granulation was sparse in both cultured cells and the original tumor . These studies demonstrate long term viability, in monolayer culture, of cells derived from this islet cell adenoma, with retention of secretory characteristics consistent with data obtained prior to removal of the adenoma from the patient.

Biochem J, 1975 Dec, 151(3), 665 - 69
Rabbit collagenase . Immunological identity of the enzymes released from cells and tissues in normal and pathological conditions; Werb Z et al.; 1 . The immunological cross-reactivity between rabbit collagenases from a variety of normal and pathological sources was examined . The specific antibody raised against collagenase secreted from normal rabbit synovial fibroblasts gave reactions of complete identity with collagenases secreted from fibroblasts derived from rabbit skin, and from synovium from experimentally arthritic rabbits . 2 . The rabbit fibroblast collagenase was immunologically identical with collagenases obtained from the organ culture medium of normal rabbit skin, synovium, ear fibrocartilage and subchondral bone . 3 . Collagenases from the culture media of normal rabbit synovium and from hyperplastic synovium of rabbits made experimentally arthritic were identical . 4 . The collagenase secreted from rabbit fibroblasts gave a reaction completely identical with that of a collagenase extracted directly from a rabbit carcinoma . 5 . IgG (immunoglobulin G) from a specific antiserum to rabbit fibroblast collagenase was a potent inhibitor of the collagenases obtained from the culture media of the various rabbit cells and tissues . 6 . Collagenases from human synovium and from mouse macrophages and bone were neither precipitated nor inhibited by antibodies to rabbit collagenase . 7 . No immunoreactive material was found in lysates of rabbit polymorphonuclear leucocyte granules with the specific antisera to rabbit fibroblast collagenase . No evidence for inactive forms of rabbit collagenase in lysates of the rabbit synovial fibroblasts could be found, either by double immunodiffusion against the specific collagenase, or by displacement of active enzyme from inhibition by the IgG.

Calcif Tissue Res, 1975 Nov 24, 19(1), 1 - 7
Net uptake and release of calcium and phosphate by bone in vitro: effects of medium calcium and phosphate concentrations; Messer HH et al.; The effects of varying the initial calcium and phosphate concentrations of the culture media on bone calcium and phosphate release were examined, using whole calvaria from 3-day-old mice in 48-hour cultures . The initial calcium and phosphate concentrations of the culture media were varied in the range 3-10 mg/100 ml; either calcium or phosphate alone was changed while the other ion was held constant, or the concentrations of both were varied while the Ca:P ratio was held constant . For all combinations, 3 treatment groups were used: i) control (no added hormone); ii) 0.5 U/ml PTH; iii) 50 mU/ml CT . The release of calcium and phosphate from the bones was greatest at low initial calcium or phosphate concentrations in the media, and least at high initial concentrations . High concentrations of both ions together abolished hormonal responses and resulted in extensive uptake of calcium and phosphate by the bones . The response to PTH was lost at a high concentration of either ion alone, while a response to CT was observed under all experimental conditions except simultaneously high calcium and phosphate concentrations.

Cell Tissue Res, 1975 Nov 12, 163(3), 365 - 72
Variability of the effects of serum-free medium, dibutyryl-cyclic AMP or theophylline on the morphology of cultured new-born rat astroblasts; Moonen G et al.; The effects of serum deprivation, of dibutyryl-cyclic AMP (dBcAMP) and of theophylline on the morphology of cultured new-born rat astroblasts have been studied using Eagle's basal medium (BME) or Eagle's minimum essential medium (MEM) as culture media . Serum deprivation had no effect on cells cultured in BME, while in MEM, deprivation induced a rapid morphological transformation involving the appearance of multiple processes . This phenomenon was rapidly reversible when serum was again added . In serum-supplemented BME, dB-cAMP (1 mM) and theophylline (1 mM) had no effect . In serum-supplement MEM, theophylline (1 mM) had no effect while dB-cAMP (1 mM) induced a slower and poorly reversible morphological alteration . On the other hand cells in serum-free BME showed multiple processes after addition of dB-cAMP (1 mM) or theophylline (1 mM) . This rapid alteration was completely reversed either by removal of dB-cAMP and theophylline or by addition of serum.

Biull Eksp Biol Med, 1975 Nov, 80(11), 36 - 8
{Dynamics of the amino acid composition of the medium during cultivation of isolated liver and kidney by the controlled perfusion method}; Barashkov VA et al.; The dynamics of the medium amino acid composition was studied during a 6-hour perfusion of dog kidney and liver by a mixture of autogenic plasma and medium 199 (a ratio of 2 : 3) . In comparison to the initial level the amount of histidine during 6-hour cultivation of the kidney was found to increase 2.2 times, that of glutamic acid--1.7 times, and of alanine and lysine--1.6 times . At the same time the amount of arginine, serine and asparagic acid became 3.3 times and of glutamine and threonine--2.5 times lower . With the liver perfusion the level of glutaminic acid became 2.9 times, of alanine--2.3 times, of cystine--2 times and of glycine--1.5 times higher . With the liver perfusion the level of arginine decreased so rapidly that none was found in the medium after a 2-hour perfusion . The described method of amino acid analysis during the cultivation of the organs could serve as means for elaborating and correcting the culture media.

Br J Dermatol, 1975 Nov, 93(5), 557 - 61
Nickel hypersensitivity . Nickel binding to amino acids and lymphocytes; Hutchinson F et al.; The nature of a protein hapten conjugate which could effect lymphocyte transformation in nickel hypersensitivity was investigated by subjecting the culture media employed to radiochromatographic analysis . Autoradiographic studies with 63Ni demonstrate direct binding of nickel salts to the lymphocyte cell surface . This occurs with a proportion of lymphocytes both from nickel sensitive and from control subjects and so binding is not of itself the stimulus for transformation.

Cancer Res, 1975 Nov, 35(11 Pt 1), 3048 - 50
The release of high-molecular-weight alkaline phosphatase and leucine aminopeptidase into the media of cultured human cells; Singer RM et al.; Using exclusion from Sepharose 4B as our criterion, we have found a high-molecular-weight form of alkaline phosphatase and of leucine aminopeptidase which are released into the culture media by the FL amnion cell line . A low-molecular-weight form of leucine aminopeptidase is also found to contribute to the total levels of this enzyme in the media . The levels of these enzymes increased during the growth cycle of the culture, paralleling the increase in cell density, suggesting that the two events may be related . This phenomen in culture suggests a possible explanation for the appearance of similar enzyme forms in patient serum and fluids originating from diseased tissue.

Clin Orthop, 1975 Nov-Dec, (113), 164 - 7
Establishment and alkaline phosphatase activity of clonal cell lines of murine osteosarcomas . A preliminary study; Amitani K et al.; Clonal cell lines of murine osteosarcomas were established and have been maintained in vitro for over a year . By implanting these cultured cells into mice, osteosarcomas, whose histological picutres were exactly the same as those of the original tumors, were easily reproduced . The cultured cells of murine osteosarcomas contain an extremely high level of alkaline phosphatase activity . The cells also secrete a great amount of extracellular alkaline phosphatase in culture media.

Clin Orthop, 1975 Oct, (112), 340 - 8
Hormone suppression of DNA synthesis in cultured chondrocyte and osteosarcoma cell line; Scranton PE Jr et al.; Recent evidence suggests that endocrine factors play an important role in the natural history of osteosarcoma . The occurrence of this tumor in the metaphysis of rapidly growing adolescents, coupled with increased female survival led to the investigation of the effects of various hormones on cultured osteosarcoma cells . The in vitro effects of physiologic concentrations of human growth hormone, 17beta estradiol, and progesterone on cultured osteosarcoma cells and chondrocytes are presented . Growth hormone significantly enhances 3H-thymidine incorporation in osteosarcoma cells and chondrocytes, in the presence of human serum . The use of other sera, culture media, or heat inactivation of the human serum abolishes this effect . Estradiol and progesterone, alone, or in combination produce significant suppression of DNA synthesis in cultured tumor cells . Several sera contain a heat-labile factor which has the capacity to block the suppressive effect of estradiol . This factor could be overcome by increasing the concentration of hormone, or by heat-inactivation of the serum . The use of hormone therapy in the treatment of osteosarcoma has never been reported, despite its demonstrated value in certain other malignancies . In light of these observations and considering the poor prognosis in this disease it seems reasonable to initiate a study of adjunctive hormone therapy in osteosarcama.

Clin Exp Immunol, 1975 Oct, 22(1), 153 - 66
An absolute requirement for serum macromolecules in phytohaemagglutinin-induced human lymphocyte DNA synthesis; Yachnin S et al.; We have examined the effect of different variables such as tissue culture media, with or without various supplements, lymphocyte isolation techniques, lymphocyte contamination by autologous red blood cells and platelets, and lymphocyte numbers, on the requirement for serum during phytohaemagglutinin (PHA) induced DNA synthesis in human lymphocytes . At all mitogen doses tested, we have found that dialysable constituents of serum enrich the ability of all tissue culture media to support lymphocyte DNA synthesis; however, human lymphocytes display an absolute requirement for nondialysable macromolecular constituents of serum in order to synthesize DNA.

Parasitology, 1975 Oct, 71(2), 311 - 26
Utilization of amino acids by Trypanosoma brucei in culture: L-threonine as a precursor for acetate; Cross GA et al.; The amino acid compositions of several culture media have been analysed and compared . The utilization and excretion of amino acids and other metabolites have been followed during growth of Trypanosoma brucei S42 in a defined medium . All of the added L-threonine was metabolized by the cells, even when it was present at elevated concentrations . Glucose was consumed throughout the growth cycle: glutamine was consumed more rapidly than glutamic acid, which was itself used at about the same rate as proline . Threonine was cleaved to form glycine and acetate, both of which accumulated in the medium . Alanine and succinate were excreted together with a small amount of pyruvate, but these three products accounted for less than half of the glucose used . CO2 production from glucose was not measured, but insignificant amounts of CO2 were produced from threonine . Tetraethylthiuram disulphide blocked the cleavage of threonine and was a potent inhibitor of trypanosome growth.

Parasitology, 1975 Oct, 71(2), 261 - 73
The measurement of the relative turnover rates of proteins of the surface membranes and other fractions of Schistosoma mansoni in culture; Kusel JR et al.; The double isotope labelling method of Arias, Doyle & Schimke (1969) and Dehlinger & Schimke (1971) was used to determine the relative rates of turnover of proteins in various tissue fractions and in the culture medium of adult Schistosoma mansoni . The majority of proteins in the surface membrane turned over at about the same rate . This is consistent with a model for membrane assembly and degradation involving multi-protein units . The proteins in the other fractions examined, except frozen-thawed supernatant fluid, also turned over at about the same rate . In the frozen and thawed supernatant fluid, the higher molecular weight (greater than 40 000 daltons) proteins turned over at a much greater rate than the lower molecular weight proteins (less than 40 000 daltons) . The antigens in the culture medium, both particulate and soluble, had a higher turnover rate than the worm tissue proteins . It is concluded that the culture medium antigens are released as the result of two distinct processes: (a) surface membrane turnover and (b) a rapid secretory process . It is possible that the high molecular weight proteins found in the forzen-thawed supernatant fluid are involved in the rapid secretory process . The culture media of 6-day and 16-day schistosomula were also examined by the double isotope method . The culture medium of the adult worm and 16-day schistosomula contains more material presumed to be secreted than does that from the 6-day schistosomula . This may explain the poor immunogenicity of young schistosomula.

J Natl Cancer Inst, 1975 Oct, 55(4), 843 - 9
Dynamic alterations in some surface properties of freshly explanted Moloney lymphoma cells; Ran M et al.; When Moloney lymphoma (YAC) cells were freshly explanted from the tumor-bearing host into culture, two events occurred in the first 120 minutes: The cells lost their natural IgG coat, and their sensitivity to complement-dependent lysis (CdL) mediated by antibodies to Moloney lymphoma cells decreased or increased . An increasing sensitivity to CdL as a function of incubation time at 37 degrees C was likely to occur when the sensitivity to CdL was low at explantation . A decreasing sensitivity to CdL was probable in instances of a high sensitivity to CdL at explantation . Artifical coating of YAC cells with antibodies to Moloney lymphoma immediately after explantation moderated the alterations in their sensitivity to CdL . This occurred even though a functional antibody did not remain on the cells as evidenced from the gradual decreased sensitivity of these artifically coated cells to the addition of complement . Spent culture media in which freshly explanted cells grew for 60 or 120 minutes sometimes blocked CdL of YAC cells mediated by antibodies to Moloney lymphoma.

Parasitology, 1975 Oct, 71(2), 247 - 59
The release of membrane antigens into culture by adult Schistosoma mansoni; Kusel JR et al.; Antigens sharing determinants with surface membranes and soluble proteins of adult Schistosoma mansoni have been detected in culture media after incubation of radioactively labelled worms . The relative quantities of these antigens were measured with specific antisera raised in rabbits and with serum from an immune rhesus monkey . It was found that 12-16% of TCA-precipitable radioactivity in the culture medium consisted of membrane antigens and 6-8% consisted of antigens sharing determinants with proteins found in the soluble fraction of adult worms . Over half the membrane antigens were present in particulate form, while other antigens were present in solution . Surface labelling the adult worms with {125I}confirmed that some of the particles in the culture medium were derived from the surface membrane of the adult worm and electron microscope examination of such particles showed that large membrane fragments were present . These results support the hypothesis that antibodies against schistosome membrane antigens are induced by particulate membrane antigens released by the parasite.

Am J Clin Pathol, 1975 Sep, 64(3), 345 - 50
Granulomatous synovitis and osteitis caused by Sporothrix schenckii; Marrocco GR et al.; Sporotrichosis must be considered in the differential diagnosis of granulomatous inflammation involving bones and joints . The organisms are difficult to demonstrate in direct smears and in histiologic sections, but they grow readily on routine fungal culture media . The cases of two patients, one with sporothrix arthritis and one with sporothrix arthritis and osteitis, are presented . The latter patient underwent ten surgical procedures over a period of 6 1/2 years and was treated for tuberculous arthritis without a definite diagnosis before fungal cultures were obtained and Sporothrix schenckii isolated.

J Bacteriol, 1975 Sep, 123(3), 992 - 9
Regulation of glutaminase levels in Escherichia coli; Prusiner S; Nitrogenous metabolites, cyclic adenosine 3':5'-monophosphate (cAMP), and the stage of culture growth all influence the levels of glutaminase A in Escherichia coli, but no variables in culture conditions alter the levels of glutaminase B . Growth of E . coli on culture media containing glucose and excess ammonia results in a rise in the level of glutaminase A as the cultures enter stationary phase; this rise is abolished by ammonia limitation . cAMP or glycerol reduce the level of glutaminase A . In mutants deficient in cAMP receptor protein, glutaminase A levels are unchanged by cAMP, but they are still susceptible to regulation by ammonia . We consider glutaminase B to be a constitutive enzyme, since its levels appear independent of nutritional conditions.

Br J Haematol, 1975 Aug, 30(4), 425 - 34
Ferritin synthesis in lymphocytes, polymorphs and monocytes; Summers M et al.; A rapid two-stage method has been devised for the separation of different leucocyte populations from human blood . Different cell types can be obtained in an undamaged state and with little contamination . Ferritin and total protein synthesis has been determined by measuring {14C}leucine incorporation in culture media which contain varying amounts of added ferric iron or desferrioxamine . Both ferritin and total protein synthesis is greater in monocytes than in lymphocytes or polymorphs when the basal medium is used . Only monocytes show a consistent increase in ferritin production due to iron stimulation . Ferritin synthesis by monocytes, polymorphs and lymphocytes is inhibited in the presence of desferrioxamine.

Cancer Res, 1975 Aug, 35(8), 1950 - 6
Effect of methotrexate on thymidylate synthetase in cultured parenchymal cells isolated from regenerating rat liver; Bonney RJ et al.; The effect of methotrexate (MTX) on thymidylate synthetase activity during liver regeneration was examined with parenchymal cells isolated 22 and 44 hr after partial hepatectomy and cultured as a monolayer . The synthetase activity in these cells decreased with a half-life of 18 to 24 hr, but if MTX (1.5 X 10(-6) to 1.5 x 10(-5) M) was present in the culture media, this decline could be delayed for at least 48 hr . In contrast, thymidine kinase activity decreased at a rate which we unaffected by MTX . Dihydrofolate reductase was inhibited at all concentrations of MTX used to block the decrease in synthetase activity . Folic acid at 10(-4) M, although less effective than MTX, also delayed the decrease in synthetase activity . The addition of cycloheximide, puromycin, or antinomycin D to the culture media did not alter the response of the synthetase to MTX . The latter studies, coupled with those indicating that the rapid loss of synthetase activity in crude extracts could be prevented by MTX or, more effectively, by MTX plus deoxyuridine 5'-monophosphate, suggest that the primary effect of MTX on thymidylate synthetase in vivo is that of enzyme stabilization . Similar stabilizing effects were obtained in liver cell extracts with 10(-5) M deoxyuridine 5'-monophosphate in combination with 10(-4) M folate or 10(-4) M dihydrofolate.

J Natl Cancer Inst, 1975 Aug, 55(2), 339 - 43
Inhibitory effect of adult bovine serum on release of infectious Epstein-Barr virus from a virus-carrier cell line; Sairenji T et al.; When certain bovine sera (BS) were used to culture P3HR-1 cells, the amount of infectious Epstein-Barr virus (EBV) released into the culture fluid was significantly less than that obtained when calf serum (CS) or fetal calf serum (FCS) was used . A direct dose-response relationship was noted between the concentration of BS and the degree of inhibition of virus production . Cell growth was not inhibited nor was virus infectivity neutralized by BS . Similarly, EBV infectivity was not reduced by spent medium containing BS . The inhibition of virus production by BS could be reversed by replacement of the serum supplement with CS . The formation of viral capsid antigen and of infectious virus within the cells was not inhibited by BS, whereas the amount of virus released was significantly decreased in comparison with culture media containing FCS or CS . These findings suggested the presence of a factor(s) in the BS which inhibited the release of EBV from the cell.

Prostaglandins, 1975 Jul, 10(1), 67 - 85
Prostaglandin synthesis by rheumatoid synovium and its stimulation by colchicine; Robinson DR et al.; The synthesis of prostaglandins by rheumatoid synovial tissue in organ culture was studied utilizing radioimmunoassay, with antisera to PGB1, PGF1alpha and PGF2alpha . It was established that PGE2 and PGF2alpha were the major prostaglandins formed by analyses of culture media with the two antisera to PGF, before and after alkali treatment . Indomethacin at 5 mug/ml suppressed prostaglandin synthesis, usually to less than 1% of control cultures . Colchicine, 0.1 mug/ml resulted in marked stimulation of prostaglandin synthesis, in some cases over 10 fold . It is suggested, because of the colchicine effect, that the state of the microtubules may regulate the rate of prostaglandin biosynthesis . It is possible that prostaglandin E2 produced by rheumatoid synovia may contribute to the pathogenesis of the inflammatory reaction and lead to destruction of juxta-articular bone in rheumatoid arthritis.

Can J Microbiol, 1975 Jul, 21(7), 1032 - 40
The recovery of nocardial recombinants from broth cultures; Brownell GH et al.; Broth culture media were examined for their ability to support growth and recombination between compatible strains of Nocardia erythropolis . Nutrient(Nut) and peptone-yeast-extract (PY) broths supported the production of recombinants after 36 h of incubation with a maximum recovery of about 6.0 times 10(-7) CFU/ml . Cells mated in trypticase broth (TB) yielded the highest incidence of recombinants (1.0 times 10(-2) CFU/ml) in the absence of parental cell growth . From a chemically defined mating broth (CD), supplemented with limited amounts of the parental-growth requirements, recombinant recovery reached about 1.0 times 10(-4) after 120 h of incubation . The recombinant class types obtained from Nut- or PT-mated strains were predominantly auxotrophic while TB-mated strains produced stable proteotrophs . The high incidence of recombinant types from TB-mated strains was due to growth of selected prototrophic classes . Studies with strains mated in PY broth indicated that the mating event occurs at very low frequencies between older, stationary-phase cells rather than between actively growing, log-phase cells.

J Natl Cancer Inst, 1975 Jul, 55(1), 7 - 10
Production of ring-shaped particles by normal and metaplastic tissue . I . Human skin; Rounds DE et al.; Maintenance media incubated with biopsy specimens of human skin tissues contained minute (10-12 nm wide), ring-shaped particles (RSP) similar to those described previously in culture media of mammalian cell lines . In addition to the qualitative demonstration of the particles by electron microscopy, a quantitative method was applied to estimate in media the amount of DNA that could be attributed primarily to RSP content . The amounts of DNA, obtained with 146 test specimens, varied with the pathologic condition of the tissue in the following ascending order: normal skin, verruca vulgaris, seborrheic verruca, actinic keratosis, basal cell carcinoma, and squamous cell carcinoma.

J Immunol Methods, 1975 Jul, 7(4), 387 - 92
Non-specific interference of certain components of tissue culture media with the radioimmunoassay of rat alpha-foetoprotein; Dambuyant C et al.; Interferences of 'Williams' tissue culture medium, used for cultivating rat hepatocytes, upon rat alpha-foetoprotein (AFP) radioimmunoassay have been investigated; they are not due to foetal calf serum proteins which are added as growth factor and can be abolished by dialysis which appears to be necessary for the distinction between AFP non-producer and low-producer cell lines . Of the three major groups of non-mineral components examined, amino acid solution played a major role; when individual amino acids were examined using the double antibody technique, arginine was found to interfere predominantly; its dose-response curve was parallel to that of rat AFP, which confirmed that an immunological identity between two substances cannot be established on the basis of parallelism as the only criterion.

C R Acad Sci Hebd Seances Acad Sci D, 1975 Jun 16, 280(23), 2685 - 8
{Metabolism of pregnenolone 16-3H and progesterone 4-14C by 18 day fetal mouse gonads in organotypic culture . Effect of the gonadotropic hormone LH}; Pointis G et al.; Fetal mice testes convert pregnenolone-16-3H and progesterone-4-14C to testosterone in organ culture . The 3H/14C ratio in progesterone and testosterone fractions isolated from culture media suggests the importance of the delta5-3 beta hydroxysteroid pathway in our experimental conditions . LH decreases radioactive testosterone production and increases the activity of the kelta4-3-ketosteroid pathway.

Mycopathologia, 1975 Jun 14, 55(3), 169 - 73
Growth responses of two phytopathogenic fungi to fernasan in culture media; Abdalla MH; The toxic effect of fernasan (containing 25% thiram) was tested on Rhizoctonia solani and Fusarium solani in liquid and agar media . The fungicide was more effective in liquid (at 100 p.p.m.) than in solid media, where 400 p.p.m . checked the growth of R . solani, but was ineffective against F . solani . Both fungi exhibited a zone of inversion growth in liquid culture whereby certain intermediate concentrations were less inhibitory than lower or subsequently higher doses . Similar reaction was recorded for F . solani alone in fungicide-containing agar media . Sclerotium formation by R . solani was highly significant, in relation to controls, at 40 p.p.m . The abundance of chlamydospores of F . solani was coupled with cessation of conidium formation increasing fernasan doses.

Rev Epidemiol Med Soc Sante Publique, 1975 Jun-Aug, 23(4-5), 269 - 76
{Atypical mycobacteria isolated from ganglions of presumed-healthy bovine and swine (author's transl)}; Garcia-Rodriguez JA et al.; A study has been carried out to detect the frequency of atypical mycobacteria in ganglions belonging to two types of animals: bovine and swine . A total of 250 samples (150 cows and 100 pigs) were studied . The samples were decontaminated by the lauryl-sulfate technique, using two mycobacteria culture media: Lowenstein-Jensen and Coletsos base . 14 pig ganglion strains were isolated (8 M . avium; 3 M . gordonae; 2 M . fortuitum; 1 M . chelonei) and 13 from the cow samples (5 M . avium, 3 M . Kansasii; 2 M . gordonae; 2 M . fortuitum; 1 M . phlei) . The most frequently isolated mycobacteria was M . avium (48.14% of total isolated strains) . This study confirms the possible importance of these animals in the epidemiology of mycobacteriosis.

Proc Soc Exp Biol Med, 1975 Jun, 149(2), 344 - 7
Fetal bovine serum: a multivariate standard; Honn KV et al.; Chemical and endocrine parameters were investigated in commercially available fetal bovine sera intended for use as culture media supplements . A high degree of serum variability was present both within and between suppliers in all major categories investigated . It is suggested that caution be employed in the interpretation of results from experiments utilizing serum supplements without specific quantitation of possible interfering or modulating factors.

Acta Endocrinol (Copenh), 1975 Jun, 79(2), 403 - 16
The influence of androgens on protein synthesis by cultured rat epididymal tubules; Blaquier JA; Rat epididymal tubules maintained in organ culture for 3 days respond to the addition of androgens to the culture media (testosterone and dihydrotestosterone 1 x 10-minus 5 m and 1 x 10-minus 7 m) with an increased incorporation of amino acids into acid-insoluble material . Significant androgenic stimulation is observed only 24 h after addition of hormone, while an inhibitory effect is found at earlier periods . The stimulation seems to be specifically produced by androgens; it is blocked by cyproterone acetate and is not elicited by oestradiol-17beta or corticosterone . The process appears to involve RNA synthesis since actinomycin D suppresses the stimulatory effect of androgen . Evidence suggests that cAMP production is not a primordial step in the response to androgen since dibutyryl cAMP did not mimick the androgenic effect, theophylline did not potentiate the response and alpha,beta-methylene ATP, which competitively inhibits adenyl cyclase, failed to alter the androgenic effect . Radioactive testosterone and dihydrotestosterone added to the culture media showed a preferential intranuclear localization as well as extensive metabolism . DHT was found to be the principal intranuclear steroid.

Jpn J Exp Med, 1975 Apr, 45(2), 133 - 8
A successful method for mass culture of the house dust mite, Dermatophagoides pteronyssinus (Troussart, 1897); Miyamoto J et al.; Comparative studies have been carried out to find out an optimum condition for efficient mass culture of Dermatophagoides pteronyssinus (Troussart, 1897), the most dominant species of the house dust mites . Among various food materials tested for culture media, the mixture composed of 2 parts of powdered laboratory animal food, 2 parts of dried yeast and 1 part of dried fish powder was found to be most fitted . The humidity content of the medium was best when adjusted to 16% beforehand and thin layer of the food medium was prepared in Roux's-bottle with a paper plug . The plug of the bottle was opened once a week and the medium was mixed thoroughly by shaking to prevent clotting . When about 15,000 mites were inoculated into 100 g of culture media, the highest yield was obtained with relative humidity regulated at 75% and the temperature at 25 degrees C after 12 weeks of incubation . Purified mites were separated from culture media by floatation with saturated NaCl solution and the average yield of the mice was 2 to 3 g in wet weight per 100 g of the food mixture.

Acta Virol, 1975 Apr, 19(2), 150 - 4
A modified plaque method for arboviruses on plastic panels; Hronovsky V et al.; Autoclavable culture media containing an increased (10---15-fold)concentration of succinate buffer permit a comparitively long-term cultivation of cells in free gas exchange with the atmosphere . Based on them, an economical technique of plaque titration of arboviruses on plastic panels with methylcellulose overlay was developed . With seven arboviruses of three different groups and three cell lines (CV-1, PS and PK), the method proved sufficiently sensitive as compared with titrations in mice or tube cell cultures and suitable for plaque reduction tests.

Tropenmed Parasitol, 1975 Mar, 26(1), 35 - 42
{Studies on the antigens of leptospires (author's transl)}; Mazzonelli J et al.; The authors show the existence of a TR (thermoresistant) antigen capable to withstand heating at 100 degrees C for 30 minutes . This antigen is present in Patoc I, serogroup Semaranga . It absorbs the corresponding immune serum at rates exceeding 99%, and appears to be the only antigen playing a part in the absorption of agglutinins . The existence of a TR antigen was demonstrated in various pathogenic serotypes . This antigen withstands heating at 80 degrees C for 10 minutes . The TR antigen absorbs antibodies of immune sera of the corresponding strains; and, in cross agglutination reactions, antibodies of the other immune sera of the studied pathogenic leptospires . the TR antigen of serotypes icterohaemorrhagiae Wijnberg, australis Ballico and pyrogenes Salinem are not able to absorb agglutinins of sera anti-Patoc I, whereas TR/Hebdomadis and TR/Pomona absorb as much as 20% . tr/hond Utrecht IV and TR/Patoc I (by absorbing their immune sera in cross agglutination) act as homologous strains with an absorption rate of 99% . The capability of absorption of TR changes according to the composition of culture media . This antigen, heated at 80 degrees C for 10 minutes and obtained pyrogenes Salinem is used for macroscopic diagnosis in slide test procedure by mixing equal proportions of antigen and serum . A positive reaction is indicated by distinct agglutination clumps.

J Clin Microbiol, 1975 Mar, 1(3), 246 - 9
Suitability of peracetic acid for sterilization of media for mycoplasma cultures; Wutzler P et al.; The utility of peracetic acid for sterilization of serum and yeast extract additions to mycoplasma medium was studied by culturing six Mycoplasma species . Culture media containing additions that had been sterilized with peracetic acid proved to be as good as filtered components . The use of 0.05 to 0.1% peracetic acid is recommended to sterilize the serum and yeast extract additions since savings in time and equipment can be accomplished.

Blood, 1975 Mar, 45(3), 321 - 34
Human chronic myelogenous leukemia cell-line with positive Philadelphia chromosome; Lozzio CB et al.; A cell-line derived from a patient with chronic myelogenous leukemia (CML) is described . The new cell-line, which has over 175 serial passanges in a 3 1/2-yr period, has the following characteristics: (1) CML cells started to proliferate actively since they were first incubated in culture media . A threefold increase in the total number of cells was observed during the first seven passages; the cell population increased by a factor of 10 to 20 every 7 days from passage 8 through 85; from 20 to 40 times from passage 86 through 150, and more than 40 times after 150 passages . (2) The majority of the nononucleated cells are undifferentiated blasts . (3) The karyotype of all the cells examined show the Philadelphia (Ph1) chromosome and a long acrocentric marker plus aneuploidy . The Giemsa-banding studies identified the Ph1 chromosome as a terminal deletion of the long arm of chromosome 22:del(22)(q12) and the long acrocentric marker as an unbalanced reciprocal translocation of one chromosome 17 and the long arm of one chromosome 15 . (4) The CML cells do not produce immunoglobulins, are free of mycoplasma, Epstein-Barr virus, and herpes-like virus particles . (5) CML cells have no alkaline phosphatase and myeloperoxidase activities and did not engulf inert particles . (6) Cultured CML cells provide a constant source of a specific antigen . This CML cell-line represents a unique source of CML cells with meaningful indicators of malignancy for clinical and experimental studies.

Eur J Biochem, 1975 Feb 21, 51(2), 429 - 35
Isohormones: molecular forms of the human chorionic somatomammotropic hormone; Belleville F et al.; The chorionic somatommaotropic hormone extracted from the human placenta exists in several molecular forms . Analytical electrophoresis in polyacrylamide gel permits separation of a highly anodic migration form, form 1, and another form migrating slightly faster than albumin, form 2 . These two forms are active as measured by radioactive immunological analysis, form 2 being about 25 times more active than form 1 . The two forms are mutually interconvertible . The two forms may also be separated by filtration on Sephadex G-50 . However, they do not differ in molecular weight, they have the same coefficient of sedimentation and the same coefficient of apparent diffusion, measured by analytic ultracentrifugation . Glutaraldehyde and 8 M urea do not modify their electrophoretic or chromatographic behavior . On the hand, the two forms differ in their tertiary structure, with the modification depending on the greater or lesser degree of oxidation of the intra-chain disulfide groups . The two forms also exist in placental culture media and the incorporation of tritiated leucine occurs preferably in form 1 . The physiological significance of the two hormone pools is not clarified.

J Clin Microbiol, 1975 Feb, 1(2), 150 - 3
Quantitative assay of dermatophyte-infected guinea pig skin scales; Sinski JT et al.; A technique to obtain homogeneous suspensions of fungal particles in scrapings from infected skin was devised that permits delivering uniform inocula to culture media being evaluated for suitability as primary isolation media for dermatophytes . The technique consisted of trypsinization of the scales, utilizing scrapings from lesions experimentally induced with an isolate of Trichophyton mentagrophytes var . granulare on guinea pigs . Optimal conditions for dislodging the fungal particles were assessed by colony counts . One-hour treatments with 0.5% (1:300) trypsin yielded the highest counts.

Parasitology, 1975 Feb, 70(1), 53 - 65
Cytology and kinetics of microgametogenesis and fertilization in Plasmodium yoelii nigeriensis; Sinden RE et al.; The sexual development of the microgametocyte of Plasmodium yoelii nigeriensis may be subdivided into microgametogenesis which includes exflagellation, dispersal of the gametes and fertilization . Under our experimental conditions microgametogenesis takes about 8-15 min at 20 degrees C, the duration of this period being inversely related to temperature . Exflagellation takes less than 1 min, subsequent dispersal of gametes may continue for 40 min . We find that exflagellation is totally inhibited in vitro by temperatures of 30 degrees C and above, and by certain invertebrate tissue culture media . Exflagellation may occur within a persistent host cell plasmalemma, which seriously impedes the escape of the 8 microgametes . Microgametes move by sinusoidal or helical waves which may be rapid (10 waves/s), slow (smaller than 1 wave/s) or they may be immobile . Microgamete activity, which is alternately rapid/slow or slow/immobile, declines linearly with time . Fertilization which takes less than 1 min results from characteristic behavioural changes by the microgamete; vibratory waves are responsible for the penetration of the macrogamete by the microgamete . The microgamete completely enters the macrogamete and therein continues its cyclical activity.

Proc Natl Acad Sci U S A, 1975 Feb, 72(2), 483 - 7
Control of ovarian cell growth in culture by serum and pituitary factors; Nishikawa K et al.; An ovarian cell line was developed that requires hormonal conditioning of the host for growth in vivo and that requires special factors for growth in vitro . It is necessary to prepare special culture media to demonstrate the effects of growth factors in vitro . To this end, methods were developed for removing from serum those essential factors required for the growth of ovarian cells in culture . Minimal growth occurred in medium containing fetal calf serum that had been passed through a porcelain filter . This method of depleting serum was replaced by a procedure involving passage through a carboxymethylcellulose column . Either pituitary extract or the eluate from the column restored growth in these depleted media . The eluate was more active than pituitary extract with regard to maximal growth enhancement . When the cells were incubated in the depleted media, viability, as measured by plating efficiency, decreased with incubation time . Either pituitary extract or the eluate from the column prevented such death of cells . Based on these findings, we postulate that the eluate contains both a survival factor and a growth-promoting factor for these ovarian cells, while pituitary extract contains only the survival factor.

Am J Pathol, 1975 Feb, 78(2), 175 - 90
Human atherosclerotic plaque cells and leiomyoma cells . Comparison of in vitro growth characteristics; Moss NS et al.; Cells derived from human atherosclerotic plaques and from arterial media were compared with cells obtained from human leiomyomata and myometrium with respect to growth behavior in long-term cell culture . None of numerous variations in culture media, including alterations of serum concentration and source, improved the rate of cell multiplication or in vitro longevity . Both uterine cell types, but neither arterial cell type, multiplied after tissue dissociation with enzymes (elastase, collagenase, hyaluronidase) . The replicative life-span of each of eight samples of arterial plaque cells was equal to or less than that of the corresponding medial cells . A similar relationship was observed for eight paired sets of leiomyoma and myometrial cells . The results indicate that, under the conditions of culture in vitro, cells of a bona fide smooth muscle tumor have a finite replicative life-span and smooth muscle cells of atherosclerotic plaques behave in a similar manner.

Neuroendocrinology, 1975, 19(2), 150 - 9
Vasotocin biosynthesis by neurohypophysial cells from human fetuses . Evidence for its ependymal origin; Pavel S; Cultured neurohypophysial cells from human fetuses aged 130-155 days release into the culture medium an active principle which has antidiuretic, hydroosmotic and rat uterine activities . The chromatographic mobility of the active principle, as well as the susceptibility of all of the above activities to tryptic digestion, indicates the presence of a basic peptide identical to arginine vasotocin (AVT) . The ability of these cultured cells, which are probably ependymal cells, to release the above activities during 43 days of incubation, suggests that AVT is synthesized and secreted by specialized ependymal cells in the developing human neurohypophysis . Neither in the culture media from anterior pituitaries of the same fetuses nor in control culture media could antidiuretic, rat uterine or hydroosmotic activities be detected . Although the nonincubated neurohypophyses of the same age contain pharmacological activities suggesting the presence of vasopressin, oxytocin and AVT, the neurohypophysial cells cultured in vitro apparently release only AVT into the medium . This supports the suggestion that whereas vasopressin and oxytocin are synthesized in the hypothalamic neurosecretory cells, AVT, on the contrary, appears to be synthesized by ependymal neurohypophysial cells in the developing human neurohypophysis . Consequently, the fetal human neurohypophysis, and presumably the fetal mammalian neurohypophysis, appears not only as a storage site for neurohypophysial hormones but also as an endocrine structure which synthesizes and secretes AVT by ependymosecretion.

Rev Asoc Argent Microbiol, 1975 Jan-Apr, 7(1), 1 - 9
{Nocardia isolated from the respiratory tract}; De Guerra CA et al.; Twelve strains of Nocardia isolated from sputa, bronchial washings and pleural fluids, were studied . The classification was based on the following characteristics: a) acid-fastness; 2) ability to disolve crystals of tyrosine and xantine; 3) hydrolysis of casein; 4) growth in dilute gelatin medium (0.4%) and 5) galactose acidification . Direct cultivation of the clinical material in Czapek liquid culture medium without carbon source and containing a paraffin rod (pariffin-bait technic), as well as the routin T.B . sputum digestion and concentration with NaOH solution followed by its cultivation in Loewenstein's medium, were employed for the isolation of Nocardia . The sensitivity of the strains to tuberculostatic drugs was investigated using the Canetti's method, as was the ability to growth in Sabouraud and lactrimel culture media supplemented with 100 mug/ml of rifamycin . Finally the pathogenicity test was carried out by intraperitoneal and intramuscular inoculation in guinea-pigs . All the strains were classified as Nocardia asteroides . Seven starins were isolated from patiens with lung conditions of no nocardial etiology . The diagnosis of nocardiosis of the lung was demonstrated in the remaining five patiens by microscopical observation of Nocardia in clinical specimens, by the repeated cultures obtained from these materials and, finally, by the fact that an improvement of the clinical symptomatology was obtained by sulfa administration.

Rheumatology, 1975, 6, 329 - 37
Mycoplasmas and 'diphtheroids' in rheumatoid arthritis; Stewart SM et al.; The failure to isolate conventional mycoplasmas from 88 synovial membranes and 119 synovial fluids from patients with proven rheumatoid arthritis using a variety of culture media and techniques agrees with the results of recent workers and suggests that these organisms are unlikely to play a role in the aetiology of the disease . In contrast, 'diphtheroid' organisms were isolated from 27 to 30% of rheumatoid specimens, but not from cultures of non-rheumatoid material . A significantly higher incidence of raised agglutination titres to C . acnes and to a 'diptheroid' isolated from a patient with rheumatoid arthritis in sera from patients with rheumatoid arthritis supports the conclusion that these organisms are present in the joints in such patients, though their role in this disease has yet to be established.

Ann Rech Vet, 1975, 6(2), 155 - 63
{Toxinogenic moulds in silage . II . -- In vitro kinetics of patulin and byssochlamic acid biosynthesis by Byssochlamys nivea Westling in liquid medium (author's transl)}; Escoula L; A strain of Byssochlamys nivea (E 70), isolated from an silage and in pure culture, produces two mycotoxins (patulin and byssochlamic acid) . A comparison of the toxinogenesis of this fungic species on eight culture media (table I) shows that maximal patulin concentration (816 p.p.m.) is obtained on Czapek's liquid medium (Difco) enriched with yeast extract (2 p . 1000) and glucose (8 p . 1 000) ; maximal byssochlamic acid concentration (390 p.p.m.) is obtained on wet corn grains (80 p . 100 of water) . The lowest amounts are found on soya meal (patulin : 30 p.p.m.; byssochamic acid : 15.2 p.p.m.) . Study of patulin and byssochlamic acid biosynthesis kinetics on liquid medium at 26 degrees C reveals that maximum patulin is obtained on day 9 of culture . It precedes maximum growth of the mould, and appears to coincide with phialospores formation . Maximum production of byssochlamic acid is obtained after 60 days of culture, that is, during mould autolysis phase . Kinetics of these mycotoxin productions should be studied in silage storage conditions.

Acta Microbiol Pol A, 1975, 8(3), 131 - 9
L-asparaginase activity of Mycobacterium phlei under various growth conditions; Pastuszak I et al.; Seven Mycobacterium strains were grown statically on salts-glycerol-asparagine (Sauton) or on salts-glucose-glutamate (Sym) media . At desired time of incubation, the bacteria were washed with water, disintegrated with powdered corundum and in resulting cell-free extracts L-asparaginase activity was determined by the Conway method . The majority of experiments were performed on M . phlei which exhibited considerable rise in L-asparaginase activity with increasing age of the culture . This change did not occur on Sym medium because of Zn2+, which proved to abolish the effect of the enzyme induction in vivo but did not inhibit the activity in vitro . Addition of rifampicin to Sauton culture media resulted in a low enzyme level . Exogenous asparagine and glycerol were not indispensable for the enzyme synthesis and could be replaced by glutamate and glucose, respectively.

Acta Microbiol Pol B, 1975, 7(2), 111 - 24
Influence of amino acids, thiourea and hydroquinone on cellulolytic activity in some Fusarium strains; Szajer C; Studies were carried out on the influence of various amino acids: cysteine, cystine, methionine, alanine, serine, tryptophan, and of thiourea and hydroquinone on the activity of cellulase synthesized by four strains of Fusarium . It follows from the results obtained that the addition of amino acids, thiourea or hydroquinone to the culture media stimulates or inhibits the activity of the enzyme studied . The effect of this action depended both on the compound and the strain studied.

C R Seances Soc Biol Fil, 1975, 169(3), 548 - 52
{Organotype culture of glandular tissue of the testis of Gabius niger L . Association with hypophysis}; Bonnin PJ; The endocrine gland of the testis of Gobius niger L . was cultured separately for three or six days by dissecting out from the seminal mass . During the sexual resting periode, the explantation don't induce histological changes; however, during the phase of high sexual activity, the explantation involves marked asthenic signs . This phenomenon is in all cases reversed by the association with pituitaries, even after a preliminary isolated culture of three days . The association with pituitaries increases also the functional activity of the explants compared with the controls tested by measuring the total amount of testosterone contained in pooled culture media, or the global metabolism of 14C sodium acetate.

J Clin Microbiol, 1975 Jan, 1(1), 44 - 9
Influence of oxygen and culture media on maintenance of whole hamster trachea organ cultures and replication of Sendai virus; Schiff LJ et al.; Effects of various oxygen concentrations and culture media on the maintenance of 4-day-old hamster trachea organ cultures and the yield of Sendai virus were studied . The basic media used were Eagle minimal essential medium, medium 199, and CMRL 1066 supplemented with glutamine and antibiotics and buffered with NaHCO3 or N-2-hydroxyethyl-piperazine-N'-2'-ethanesulfonic acid (HEPES) . In addition, each medium was evaluated under a gas phase of 5% CO2 and 95% O2, 5% CO2 and 45% O2 and 50% N2, or 5% CO2 and 95% air . Culturing of explants with CMRL 1066 and medium 199 buffered with HEPES in the presence of 5% CO2 and air proved most efficient; ciliary movement and ciliated surface epithelium were maintained for periods up to 27 days . No significant difference in the rate of replication of Sendai virus was seen in the three different media with the two buffer systems in the three different gaseous phases . The addition of 0.2% bovine serum albumin to the media yielded greater quantities of virus, up to 200-fold increase in titer without producing changes in ciliary function . A distinctive pattern of morphological changes was observed in explants of trachea epithelia inoculated with Sendai virus . These results suggest the practical application in the use of whole hamster trachea explants as a diagnostic aid in the isolation of Sendai virus from laboratory rodents.

Bibl Haematol, 1975, (40), 621 - 5
Hexose transport in sarcoma virus transformed cells; Hatanaka M et al.; Avian and mammalian fibroblast cultures transformed by type C sarcoma viruses show a dramatic enhancement of the rate of hexose transport at the beginning of transformation which is quantitatively and qualitatively different from that seen by variation in culture conditions of nontransformed control cells . The identification of this change as being a transport alteration independent of total glucose metabolism has been shown by use of nonmetabolizable analogues, 2-deoxyglucose, 3-O-methylglucose, and L-glucose . Increased transport rates were not dependent on levels of hexokinase activity . Transport studies of 3-O-methylglucose confirmed these conclusions and further revealed an additional altered nature of hexose transport after transformation by sarcoma virus . 3-O-methylglucose was not only transported more rapidly in the transformed cells than in the parental nontransformed cells, but the sugar "infiltrated" into the transformed cells despite the inhibitory effect of cytochalasin B . This was not seen with control cells . The sarcoma cells were also able to transport L-glucose in contrast to lack of uptake by nontransformed cells . Under conditions in which cell toxicity was not a factor, 2-deoxyglucose and several other sugars present in culture media inhibited transformation by sarcoma viruses . These same sugars reduced the incidence of sarcomas produced by virus in vivo when administered daily to test animals . The transport changes also correlate well with the transformed state as found by other laboratories using temperature-sensitive mutants and revertant cell lines . Collectively these data suggest that manipulation of transport systems may prove useful for control of certain malignancies.

C R Seances Soc Biol Fil, 1975, 169(6), 1429 - 35
{Influence of the oxygenation state of the culture media on the level of non esterified fatty acids and of cholesterol in heart cells of the newborn rat}; Freyss-Beguin M et al.; The uptake of non esterified fatty acids (NEFA), from a medium supplemented with 10% of calf serum, by rat myocardial cells cultured during 8 days, without renewal of the medium, was identical whatever the values of partial pressures of oxygen in the medium . Cellular cholesterol level was reduced when air or oxygen (+5% CO2) was blown into culture bottles; it was enhanced by gassing with nitrogen 24 h later . Cellular NEFA level was reduced by air, but not by oxygen blowing; it was enhanced by gassing with nitrogen 24 h later . The variations observed may originate from pertubations of NEFA beta-oxidation.

Connect Tissue Res, 1975, 3(2), 115 - 22
Stimulation of chondroitin sulfate proteoglycan production by chondrocytes in monolayer; Schwartz NB et al.; Chondrocytes in monolayer undergo morphological and biochemical changes which culminate in the establishment of cartilage nodules in vitro . Chondroitin sulfate or heparin, added to the culture media of these cells, stimulates the production of chondroitin sulfate proteoglycan over the entire period of culture with a maximum effect during the log phase of growth . In addition, a lag of 2-3 hours is required before an increase in sulfate incorporation into polysaccharide is observed . The responsiveness of chondrocytes is influenced by several factors, such as cell density, conditioned media and enzyme treatment . Furthermore, puromycin abolishes the endogenous as well as the stimulated synthesis, demonstrating the necessity for core protein synthesis in both synthetic processes . Addition of beta-D-xylosides (which presumably act as initiators of chondroitin sulfate polysaccharide synthesis) and chondroitin sulfate, concurrently, stimulate sulfate incorporation to levels higher than either agent alone, indicating that these compounds act by different mechanisms.

Can J Microbiol, 1975 Jan, 21(1), 17 - 25
Environmental factors affecting the degradation of Dyfonate by soil fungi; Flashinski SJ et al.; The ability of selected fungi to degrade the soil insecticide Dyfonate (O-ethyl S-phenyl ethylphosphonodithioate) into water-soluble, noninsecticidal metabolites was found to be dependent on the supply of nutrients, incubation time, temperature, pH, as well as other factors . With yeast extract as the carbon source (5 g/liter) and ammonium nitrate (1 g/liter) as the nitrogen source, both Rhizopus arrhizus and Penicillium notatum degraded the insecticide to a larger extent than with any other combination of nutrients used . With glucose as the carbon source, concentrations of ammonium nitrate above 5 g/liter inhibited the degradation of Dyfonate by R . arrhizus . Time-course studies on the metabolism of the insecticide indicated that Dyfonate was first absorbed by the fungal mycelium, where it was metabolized followed by the release of water-soluble, noninsecticidal, breakdown products into the culture media . The degradation appeared to involve the breakdown of Dyfonate into ethyl acetate soluble metabolites, such as ethylethoxyphosphonothioic acid, ethylethoxyphosphonic acid, methyl phenyl sulfoxide, and methyl phenyl sulfone . These compounds were then further degraded into water-soluble products . The optimum conditions for the degradation of the insecticide by R . arrhizus were observed at pH 6.0 to 7.0 and at 15-25 degrees C . Aged fungal mycelia were as active as mycelia in the logarithmic growth phase.

Proc Soc Exp Biol Med, 1971 Oct, 138(1), 98 - 102
Antiserum inhibition of rabbit spermatozoal adherence to ova; Menge AC; PIP: The in vitro effects of different normal and immune sera, rabbit oviductal fluid, and culture media from incubations of female reproductive tissues on the adherence of rabbit spermatozoa to rabbit and rat ova is described . There were no apparent differences in results due to species of ova . Adherence of spermatozoa to ova was not affected by treatment with normal sera . Antisera against materials that contained spermatozoa such as semen epididymal spermatozoa and testis completely inhibited the spermatozoa from attaching to ova and caused considerable agglutination of sperm cells in the incubation slides . When the agglutinating and immobilizing activities of rabbit and goat antisera were removed by papain treatment the antiadherent effect of the antisera was unaffected . Oviductal secretions and concentrated media from the culture of reproductive tissue from female rabbits isoimmunized with semen inhibited adherence of sperm .

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