scholarly journals A METHOD FOR THE PHYSIOLOGICAL STUDY OF TISSUES IN VITRO

1923 ◽  
Vol 38 (4) ◽  
pp. 407-418 ◽  
Author(s):  
Alexis Carrel
Keyword(s):  
Nutrient Medium ◽  
Bacterial Contamination ◽  
Culture Medium ◽  
Residual Activity ◽  
Physical Factors ◽  
Dynamic Changes ◽  
Fluid Medium ◽  
Continuous Growth ◽  
Pure Cultures

1. A method has been developed which allows the continuous growth of pure strains of fibroblasts, epithelium, and leucocytes in a medium which undergoes but slight spontaneous deterioration. 2. The principle of the method is to leave the tissues undisturbed while the medium is changed. This was realized by special containers allowing the change of the medium without bacterial contamination and by the simultaneous use of a solid and a fluid medium. 3. The curve of growth of pure cultures of fibroblasts and epithelial cells in a nutrient medium is a parabola; in a non-nutrient medium, it is S-shaped and expresses the residual activity of the tissues. Leucocytes invade the culture medium progressively, as do bacteria, but never aggregate in a tissue. 4. The method is used for the study of the morphological and dynamic changes occurring in tissues under the influence of chemical and physical factors.

scholarly journals Effective medium for in vitro sprouting of the buds and multiplication of the plantlets, and identification of the fungal contaminant associated with the explants of Gnetum

2021 ◽  
Vol 16 (2) ◽  
pp. 001-013
Author(s):  
Abwe Mercy Ngone ◽  
Lawrence Monah Ndam ◽  
Rita Mungfu Njilar ◽  
Doungous Oumar ◽  
Thomas Eku Njock
Keyword(s):  
Culture Medium ◽  
Culture Media ◽  
Fungal Growth ◽  
Growth Media ◽  
Fungal Contamination ◽  
Surface Sterilization ◽  
Pure Cultures ◽  
Nodal Cuttings ◽  
Baseline Information

Plant tissue culture requires the optimization of growth media. Gnetum, known locally in Cameroon as “Eru” is an indigenous gymnospermous vegetable with diverse medicinal, nutritional, cultural and socio-economic values. This resource is over-exploited and expected to neighboring countries, resulting to increased scarcity in the forest. Preliminary work on the in vitro culture of nodal cuttings was faced by the problem of fungal contamination. It was therefore necessary to isolate and identify the fungal contaminant, optimize the surface sterilization of field material and compose an appropriate medium for sprouting. Pure cultures of the fungus were obtained and grown on Potato Dextrose Agar (PDA) and Sabouraud Dextrose Agar (SDA). The identification was based on the appearance of the fungal growth on plates and also on the microscopic view. This was affected by the use of keys. Gnetum explants were disinfected with the various concentrations of disinfectants, preceded in some instances by pre-treatments, as well as incorporating fungicides in the culture medium. Two different culture media were employed: the Woody Plant Medium (WPM) and the Murashige and Skoog (MS) based establishment medium (Y-1). Gnetum was found to live in association with a complex of Microsporum species. The level of contamination of cultures was reduced from 100% to 40% when pre-treated before disinfection and even lower to 10% by incorporating fungicides in the medium. Sprouting was observed in WPM. This study provides baseline information on the in vitro propagation of Gnetum and thus opens up avenues for more research to be carried out in this field.

Amino acid metabolism of bovine blastocysts derived from parthenogenetically activated or in vitro fertilized oocytes

1998 ◽  
Vol 10 (3) ◽  
pp. 279 ◽  
Author(s):  
Y. G. Jung ◽  
T. Sakata ◽  
E. S. Lee ◽  
Y. Fukui
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Metabolism ◽  
Culture Medium ◽  
Acid Metabolism ◽  
Essential Amino Acids ◽  
Fluid Medium ◽  
Vitro Fertilization ◽  
Oviduct Fluid

The uptake and synthesis of 19 amino acids by fresh or frozen–thawed bovine blastocysts produced by parthenogenesis (PT) or in vitro fertilization (IVF) were compared in the present study. Fresh blastocysts, 180 h after IVF or PT activation, and frozen–thawed blastocysts, 168 h old and cultured for 12 h post-thawing, were cultured in synthetic oviduct fluid medium (SOFM) containing polyvinyl alcohol (PVA) with both essential and non-essential amino acids (EAA and NEAA, respectively) (Medium 1: M1) or SOFM containing PVA with only EAA (Medium 2: M2). In Experiment 1, when fresh or frozen–thawed PT blastocysts were cultured in M1, the uptake of glutamate (in fresh only), aspartate and arginine, and the synthesis of glutamine and alanine were significantly enhanced. In the culture with M2, serine, asparagine, glutamate, glutamine, glycine, arginine and alanine were significantly taken up. It was found that the glutamine concentrations was significantly higher (P < 0.001) in the culture medium drops containing embryos than in the drops without embryos. In Experiment 2, when PT blastocysts were cultured in M1, the uptake of aspartate and synthesis of alanine were greater (P < 0.01) than those by IVF blastocysts. When M2 was used, a significant (P < 0.01) production of serine, asparagine, glutamate, glutamine and alanine, and the uptake of arginine by PT blastocysts were observed. In Experiment 3, when IVF blastocysts were cultured in M1, fresh blastocysts depleted more aspartate and glutamate, and produced more glutamine and alanine than frozen–thawed blastocysts. When cultured in M2, frozen–thawed blastocysts depleted more threonine (P < 0.01) than fresh blastocysts. These results indicate that the uptake and synthesis of amino acids were different in fresh or frozen–thawed bovine blastocysts derived from PT or IVF. These differences in amino acid metabolism may be related to the viability of the blastocysts.

scholarly journals CULTURES OF ORGANIZED TISSUES

1922 ◽  
Vol 36 (4) ◽  
pp. 393-397 ◽  
Author(s):  
Albert Fischer
Keyword(s):  
Small Body ◽  
Chick Embryos ◽  
Malignant Cells ◽  
Intestinal Tissue ◽  
Fluid Medium ◽  
Long Period ◽  
Muscle Tissues ◽  
Pure Cultures ◽  
Cultivation In Vitro

An artificial organism, if one may so term it, composed of a complex of tissues, was cultivated for a long period of time. Small fragments of intestine from chick embryos 20 to 21 days old were placed in a suitable medium. The epithelium proliferated and completely covered the fragment of intestine after 4 to 6 days. A small body was thus formed, round or oblong in shape, surrounded by cylindrical epithelium and containing epithelial, connective, and muscle tissues, endothelium, and ameboid cells. After a month's cultivation in vitro, no necrosis had occurred. Therefore, it may be assumed that, through the intestinal epithelium, the medium supplied the intestinal tissue with sufficient nourishment. No uncontrolled proliferation took place after the epithelium bad surrounded the entire fragment. The cultivation of complex tissues will facilitate the study of the interactions of the different cells under various conditions. In some experiments, pure cultures of epithelial cells were grafted into such an "organism" without difficulty. The growth of malignant cells could be studied in the same way. When the "organism" was placed in a fluid medium, the epithelium remained normal but the stroma disappeared. It seems that plasma played an important rôle in the maintenance of the tissues in their normal condition.

scholarly journals In vitro seed germination and multiplication of Calophyllum brasiliense

2016 ◽  
Vol 36 (87) ◽  
pp. 185
Author(s):  
Sheila Susy Silveira ◽  
Juliana Degenhardt-Goldbach ◽  
Marguerite Germaine Ghislaine Quoirin
Keyword(s):  
Bacterial Contamination ◽  
Culture Medium ◽  
Germination Rate ◽  
Nodal Segment ◽  
In Vitro Germination ◽  
Fungal Contamination ◽  
Natural Reproduction ◽  
Calophyllum Brasiliense ◽  
Free Rate

Calophyllum brasiliense is a tree species with limited natural reproduction. In vitro germination may be an alternative for obtaining high-quality seedlings. Seeds were maintained in water before surface disinfestation and compared with control seeds (i.e. not immersed), without differences between treatments. HgCl2 used during surface-disinfestation reduced contamination rates of cultures. Fungal contamination was reduced with fungicide added to culture medium (23 to 6.4%), although bacterial contamination increased (24 to 36%). In another experiment, seeds were immersed in plant preservative mixture (PPM™) prior to surface disinfestation. By combining immersion for 48 h and 2 mL L-1 in culture medium, contamination was only 6%. Seeds immersion in GA3 prior to surface disinfestation reduced root formation as concentration increased. Germination rate and GSI were reduced, respectively, from 72% and 0.129 (24 h) to 60% and 0.092 (48 h) according to exposure time to GA3. After 90 days in multiplication medium containing benzylaminopurine, average number of shoots per nodal segment was 3.4. In conclusion, in vitro germination of C. brasiliense seeds is feasible in sucrose-free WPM medium and reaches a high contamination-free rate (up to 93.3%).

AMINO ACIDS AND PROTEINS IN CULTURE FILTRATES OF THE ANTHRACNOSE PATHOGEN FUNGUS COLLETOTRICHUM LINI

2020 ◽  
pp. 121-127
Author(s):  
N. V. Proletova ◽  
Keyword(s):  
Amino Acids ◽  
Culture Filtrate ◽  
Nutrient Medium ◽  
Culture Medium ◽  
Protein Composition ◽  
Entire Period ◽  
Culture Filtrates ◽  
Russian Research Institute ◽  
Russian Research

The research was carried out on the basis of laboratory biotechnologies of All-Russian research institute of flax (Tver region) in 2010–2012, 2016. The aim of the work was to determine the amino acid and protein composition of culture filtrates of the anthracnose pathogen fungus Colletotrichum lini Manns et Bolley in order to adjust the concentration of selective agent in the nutrient medium when creating in vitro new flax genotypes resistant to anthracnose. It was established that the culture filtrates of strains 527 and 608 contain such amino acids as alanine, glycine, asparagine, cysteine, threonine, aspartic acid, glutamic acid, as well as arginine in strain 527 and traces of tyrosine and lysine in strain 608. It was established that the concentration of amino acids in EC of strain 527 was significantly higher than in culture filtrate of strain 608. It was shown that the toxicity of the culture filtrate depended on the degree of aggressiveness of the anthracnose pathogen strain – culture filtrate of a strongly aggressive strain is more toxic than the culture filtrate of a weakly aggressive strain. Studies have revealed that when cultivating the fungus-causative agent of anthracnose on a nutrient medium, as the mycelium of fungus grew, the concentrations of asparagine, alanine, aspartic and glutamic acids, and glycine decreased in the culture filtrates. It was established that the change in amount of proteins happened during the entire period of cultivation of the mycelium of fungus on a liquid nutrient medium. It is shown that accumulation and content of proteins in culture filtrates of strains of different aggressiveness occurs in different ways. The more aggressive strain is (639), which is more toxic, contains and accumulates more proteins in the culture medium during the entire period of growth and development the less aggressive strain is (419).

Effects of serum or fatty acid supplementation of synthetic oviduct fluid medium on development of bovine embryosin vitro

1999 ◽  
Vol 1999 ◽  
pp. 62-62
Author(s):  
N.C. Farrar ◽  
M.E. Staines ◽  
G.J. McCallum ◽  
P. Haggarty ◽  
J.J. Robinson ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Culture Medium ◽  
Culture Media ◽  
Bovine Embryo ◽  
Fluid Medium ◽  
Alternative Approaches ◽  
Defined Culture ◽  
Oviduct Fluid ◽  
Transmission Of Disease

Serum, which is routinely included in many embryo culture media, can decrease the viability of bovine and ovine embryos produced in cultures employing synthetic oviduct fluid (SOF; Kuran et al., 1999) and represents a possible route for transmission of disease. Alternative approaches include the use of chemically defined culture media but results from studies which avoid sera and its derivatives (e.g., albumin) are generally less favourable due to a lack of knowledge regarding the embryo's response to specific nutrients, most notably fatty acids. As a preliminary step towards investigating fatty acid influences on bovine embryo developmentin vitro, the present study examined the effect of adding palmitic acid (C16:0) to SOF plus bovine serum albumin (BSA) on the performance of this semi-defined culture medium and contrasted it with embryo production in SOF supplemented with serum.

In vitro sporulation of Frankia strain HFPCcI3 from Casuarina cunninghamiana

1991 ◽  
Vol 37 (12) ◽  
pp. 897-901 ◽  
Author(s):  
Stephen H. Burleigh ◽  
Jeffrey O. Dawson
Keyword(s):  
Amino Acids ◽  
Nutrient Medium ◽  
Culture Medium ◽  
Potassium Phosphate ◽  
Sulfur Amino Acids ◽  
Nitrogen And Phosphorus ◽  
Frankia Strain ◽  
Casuarina Cunninghamiana ◽  
Low Culture

Optimization of the in vitro sporulation of Frankia is a prerequisite for the development of spore preparations as a practical inoculum for actinorhizal plants. The in vitro sporulation of Frankia strain HFPCcI3 previously isolated from Casuarina cunninghamiana was increased from 0 to 12 million spores per millilitre of culture medium when nitrogen and phosphorus were excluded from a defined nutrient medium. The same medium did not increase sporulation of Frankia isolates HFPAr13 from Alnus rubra and HFPCpI1 from Comptonia peregrina. The addition of 10 mM aliphatic L amino acids increased the in vitro sporulation of HFPCcI3 by as much as 39% when grown in a defined nutrient medium with 1 mM potassium phosphate lacking ammonium chloride, whereas the addition of 10 mM acidic, basic, aromatic, and sulfur amino acids decreased sporulation. The in vitro sporulation of HFPCcI3 was inhibited by culture at 33 °C relative to culture at 23 and 28 °C. Rotary shaking at 100 rpm increased sporulation at the low culture temperatures. Key words: actinorhizal, Casuarina, Frankia, HFPCcI3, sporulation.

scholarly journals THE DRUG-FASTNESS OF SPIROCHETES TO ARSENIC, MERCURIAL, AND IODIDE COMPOUNDS IN VITRO

1917 ◽  
Vol 25 (3) ◽  
pp. 349-362 ◽  
Author(s):  
Seinai Akatsu ◽  
Hideyo Noguchi
Keyword(s):  
Culture Media ◽  
Treponema Pallidum ◽  
Chemotherapeutic Agents ◽  
Inhibitory Influence ◽  
Fluid Medium ◽  
Rate Of Increase ◽  
Fluid Media ◽  
Pure Cultures ◽  
Free Media

In the foregoing experiments we attempted to determine whether or not, by subjecting several varieties of spirochetes to increasing doses of certain chemotherapeutic agents, a gradual increase of resistance to the latter could be shown. For this purpose, pure cultures of Treponema pallidum, Treponema microdentium, and Spirochœta refringens were used against the action of salvarsan, neosalvarsan, bichloride of mercury, and iodine-iodide potassium solution in vitro. For culture media, the usual ascites-broth-tissue medium as well as solid ascites-agar-tissue medium was used. After permitting the spirochetes to grow for a fortnight in media containing certain quantities of each drug, transfers were made from tubes showing various degrees of growth to the next series of tubes containing the same drug in still higher concentrations, and similar transfers repeated every 2 weeks. The results of the experiments may be briefly summarized as follows: 1. Treponema pallidum and Treponema microdentium have, within 3 to 4 months, increased their tolerance to salvarsan and neosalvarsan to five and one-half times their original mark. With Spirochata refringens the increase was about three times. 2. Against the action of bichloride of mercury, the amount of increased tolerance of Treponema pallidum was about 35 to 70 times the original, while that of Treponema microdentium was about 10 times as much and was reached within 10 weeks. Spirochata refringens resisted 30 times the original dose. 3. There was an unmistakable increase of resistance of these spirochetes to the action of the iodine-iodide solution (Lugol's solution) when they were grown for several generations in fluid media containing the iodine solution, but the rate of increase between the initial and the acquired tolerance was slight. In general, the addition of Lugol"s solution to fluid media has a weak inhibitory influence upon the growth of the spirochetes, requiring for the total suppression of growth a quantity of over 0.7 cc. to 5 cc. of the culture media. The tolerance reached was for about three times that amount. 4. A similar tolerance phenomenon has not been established when employing a solid instead of a fluid medium containing the drugs. No explanation is offered except a suggestion that the drugs held in the agar do not enter into combination with certain tissue constituents of the medium as they are able to do with tissue elements in fluid media. This may be a factor necessary for inducing drug tolerance in these organisms in vitro. 5. The increased drug-fastness in vitro has a limit beyond which no further advance can be made. This limit varies with different species of spirochetes. 6. The acquired drug-fastness in vitro gradually disappears when the spirochetes are cultivated again in the drug-free media for several generations.

scholarly journals Role of TGFβ1 and WNT6 in FGF2 and BMP4-driven endothelial differentiation of murine embryonic stem cells

Angiogenesis ◽  
2021 ◽  
Author(s):  
Anna Gualandris ◽  
Alessio Noghero ◽  
Davide Cora’ ◽  
Elena Astanina ◽  
Marco Arese ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Culture Medium ◽  
Expression Profiles ◽  
Es Cells ◽  
Embryonic Stem ◽  
Endothelial Differentiation ◽  
Loss Of Function ◽  
Pure Cultures

AbstractEmbryonic stem cells (ES) are a valuable source of endothelial cells. By co-culturing ES cells with the stromal PA6 cells, the endothelial commitment can be achieved by adding exogenous FGF2 or BMP4. In this work, the molecular pathways that direct the differentiation of ES cells toward endothelium in response to FGF2 are evaluated and compared to those activated by BMP4. To this purpose the genes expression profiles of both ES/PA6 co-cultures and of pure cultures of PA6 cells were obtained by microarray technique at different time points. The bioinformatics processing of the data indicated TGFβ1 as the most represented upstream regulator in FGF2-induced endothelial commitment while WNT pathway as the most represented in BMP4-activated endothelial differentiation. Loss of function experiments were performed to validate the importance of TGFβ1 and WNT6 respectively in FGF2 and BMP4-induced endothelial differentiation. The loss of TGFβ1 expression significantly impaired the accomplishment of the endothelial commitment unless exogenous recombinant TGFβ1 was added to the culture medium. Similarly, silencing WNT6 expression partially affected the endothelial differentiation of the ES cells upon BMP4 stimulation. Such dysfunction was recovered by the addition of recombinant WNT6 to the culture medium. The ES/PA6 co-culture system recreates an in vitro complete microenvironment in which endothelial commitment is accomplished in response to alternative signals through different mechanisms. Given the importance of WNT and TGFβ1 in mediating the crosstalk between tumor and stromal cells this work adds new insights in the mechanism of tumor angiogenesis and of its possible inhibition.

scholarly journals Designing a Nutrient Medium for the Accumulation of Microbial Mass of Listeria

2020 ◽  
Vol 5 (4) ◽  
pp. 60-66
Author(s):  
N. M. Khaptanova ◽  
S. V. Lukyanova ◽  
V. I. Kuznetsov ◽  
N. G. Gefan ◽  
N. M. Аndreevskaya ◽  
...  
Keyword(s):  
Nutrient Medium ◽  
Culture Medium ◽  
Specific Activity ◽  
Germination Rate ◽  
Biochemical Properties ◽  
Quantitative Composition ◽  
Nutrient Media ◽  
Microbiological Methods ◽  
Microbial Mass

Background. To obtain reliable results of laboratory studies on the identification of Listeria, the presence of certified diagnostic agglutinating Listeria sera is required. An important step in the manufacturing process of such medical devices for in vitro diagnostics requires effective nutrient media for the accumulation of listeriosis microbe. Aim of the research. To develop an effective nutrient medium for the accumulation of bacterial mass of Listeria. Materials and methods. The object of the study was an experimental culture medium for Listeria cultivation. As a control, we used nutrient agar for the cultivation of microorganisms (fish meal hydrolysate, FMH-agar) and meat-peptone agar with 1 % glucose (MPA with 1 % glucose). The specific activity of nutrient media during cultivation of the test strain Listeria monocytogenes 766 was evaluated using a complex of microbiological methods. Results. The optimal base of the nutrient medium for Listeria cultivation has been selected: pancreatic hydrolysate of river magpie fish (Rutilus rutilus lacustris) and hydrolysate of meat water production waste. The qualitative and quantitative composition of the nutrient medium has been developed, its physical, chemical and biological properties have been studied. It was found that after 24 hours of incubation at 37 °C, the nutrient medium provided the growth of typical Listeria colonies. The germination rate was 85 %, which is higher compared to the growth of the culture on MPA with 1 % glucose and GRM agar by an average of 21 % (p < 0,05). Conclusion. The experimental culture medium for Listeria cultivation provided growth of colonies of the test strain L. monocytogenes 766 with the preservation of characteristic cultural, morphological and biochemical properties, and, in terms of germination and growth rate, exceeded the control media. The developed nutrient medium provides effective growth of Listeria and can be used as a medium for the accumulation of microbial mass.

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