共查询到20条相似文献,搜索用时 15 毫秒
1.
William A. Corrigall Stanley M. Crain Murray B. Bornstein 《Developmental neurobiology》1976,7(6):521-536
Explants of 18- or 19-day fetal mouse olfactory bulb have been maintained in culture for periods up to 5 weeks. Compound action potentials can be evoked in the bulb explants by 1 day in vitro, and by 3–4 days, synaptically mediated slow wave discharges can be demonstrated in bicuculline (10?5 M). The capability of the bulb explants to generate these slow-wave discharges has also been revealed by the introduction of picrotoxin (10?5 M), d-tubocurarine (10?4 M) and chloride-free medium, but not of strychnine (up to 3 × 10?5 M). The data indicate early functional development of inhibitory, as well as excitatory, synaptic systems. In addition, the selective and reversible depression of these slow wave potentials by GABA (1–5 × 10?4 M), but not by glycine (up to 3 × 10?3 M), indicates a GABA-ergic component in the inhibitory network. Single unit extracellular recordings have been obtained from the presumptive mitral cells which, in culture, are spontaneously active even as early as 1–2 days after explantation. Correlative Bodian silver-impregnations demonstrate the presence of neurons in these explants which resemble typical mitral cells. Studies of mitral cells using paired stimuli suggest the development in vitro of an inhibitory system analogous to that known to suppress the excitability of their in situ counterparts following orthodromic or antidromic activation. These data, as well as the pharmacological sensitivities of the mitral cells in culture to GABA (5 × 10?4 M) and bicuculline (10?5 M), indicate that granule-to-mitral synapses may develop characteristic functions in olfactory bulb explants. 相似文献
2.
MONA A. ABDALLAH SAMI K. ABDEL-HAFEZ FADWA M. AL-YAMAN 《The Journal of eukaryotic microbiology》1990,37(2):113-117
The continuous culturing of Trypanosoma acomys in the presence of a murine areolar-adipose cell line (A9) was possible for the 1st time. The trypanosomes were cultured at 37° C with A9 in DMEM supplemented with 20% heat inactivated fetal bovine serum, using an initial inoculum from primary cultures of lung or blood clots from infected spiny mice. The cultures were maintained for 115 days and underwent 15 passages before termination and cryopreservation. Using this culture system T. acomys subcultures were initiated from 3 different initial inocula (3 × 104, 1.5 × 105 and 7.4 × 105 parasites/ml) and growth curves revealed that the lowest inoculum gave the best growth pattern. This inoculum yielded a population doubling time of less than 12 h for 4 days, a high peak density of 7 × 106 parasites/ml and the most gradual decline compared to the other 2 inocula. Rosetting epimastigotes and nests of amastigotes were observed in close association with the feeder layer cells. Epimastigotes were the most predominant form in culture supernatants but other morphological forms observed included trypomastigotes and sphaeromastigotes. 相似文献
3.
A synchronization process is described. The lengths of the dark and light periods of each cycle are determined by the evolution
of the culture. In this way 15-litre axenic cultures are obtained. After the first cycle they contain at least 90×109
Chlorella cells. 相似文献
4.
Bjrn Holmstrm 《Biotechnology and bioengineering》1968,10(3):373-384
A HeLa cell line was propagated in semicontinuous suspension culture, 85 liters final volume, and in continuous flow culture with a volume of 300 ml. or 5 liters in an autoclavable medium to which 8% calf serum had been added. A medium containing 0.1% Methocel and 2% calf serum was also tested. Maximum productivity was obtained at a dilution rate of 0.33 day?1 with a cell density of about 1.0 × 106 cells/ml. The same cell line was also infected with Rubella virus and the production of virus was followed at the 5-liter cultivation level. 相似文献
5.
Y. Pu I. A. Bernstein L. I. Bernstam R. L. Bronaugh 《In vitro cellular & developmental biology. Animal》1995,31(4):283-287
Summary The culture of cutaneous keratinocytes grown on a Puropore nylon microporous membrane at the air-liquid interface has been
shown to be similar to the epidermis in a number of molecular and morphologic characteristics but to exhibit a significantly
greater degree of tritiated water permeation. Various culture conditions have been altered in an effort to improve the water
barrier properties. A Kp value in the range of 5.5±1.6×10−3 has been obtained for 79% of the culturea) by plating 0.9×106 viable basal cells on a piece (13-mm diameter) of membrane for 7 days of submerged growth,b) by placing two membranes on two stacked glass fiber filters (47-mm extra-thick) in a culture dish (60 mm) for 14 days of
growth at the air-liquid interface,c) by replacing the growth medium, i.e., 1 ml of complete minimum essential medium (CMEM) every 24 h after lifting,d) by using 10% fetal bovine serum (FBS) in the CMEM during the submerged culture period and 15% FBS in the CMEM during the
lifted culture period, ande) by adding a dialysis membrane on top and a Puropore nylon membrane below the culture when the cultures were inserted in
the permeation cell for testing. The percentage of cultures with this value for Kp can be increased to 90% if only cultures
with yellow, smooth, and shiny surfaces are tested. This system should be useful as a replacement for skin in testing the
cutaneous permeation of some chemicals.
To whom correspondence should be addressed at 1528 Public Health, The University of Michigan, Ann Arbor, Michigan 48109-2029. 相似文献
6.
Jianyong Wu Glenn A. King Andrew J. Daugulis Peter Faulkner Mattheus F. A. Goosen 《Cytotechnology》1992,9(1-3):141-147
Spodoptera frugiperda (IPLB-SF-21) insect cells were grown in shake-flasks and infected with a temperature-sensitive baculovirus to express the
gene of chloramphenicol acetyl transferase (CAT) in serum-free medium (SF-900) and two serum-supplemented media (IPL-41 and
Grace's). In temperature-shift experiments (cell growth at 33°C followed by virus replication at 27°C 3–4 days later), virus
and CAT production were much poorer in the serum-free medium than in serum-supplemented media, though cell growth was virtually
the same in the different media tested. In all the three media, highest virus and CAT titers were obtained at the lowest MOI
(multiplicity of infection 0.02). This result is contrary to that obtained in constant-temperature culture (27°C for both
cell growth and virus replication). Virus and CAT production was greatly improved when the entire culture was run at constant
temperature. It appeared that infected cells were severely damaged at 33°C (6°C above the optimal 27°C), resulting in little
or no virus and protein production. As a result of these temperature-shift experiments, a larger-scale (141 air-lift bioreactor)
serum-free culture of Sf-9 insect cells was conducted at constant temperature (27°C) to produce recombinant protein (β-galactosidase).
A cell density as high as 1×107 cells.ml−1, and a β-gal concentration of up to 104,000 unit.ml−1 were achieved. 相似文献
7.
Valdis Dzelzkalns Terrence Foley Samuel I. Beale 《Archives of biochemistry and biophysics》1982,216(1):196-203
Physical and kinetic properties of δ-aminolevulinic acid synthase from wild-type and aplastidic strains of Euglena gracilis have been determined. Michaelis constants for glycine, succinyl-CoA and pyridoxal phosphate are 8.5 × 10?3m, 2.5 × 10?5m, and 2.9 × 10?6m, respectively. Optimum reaction pH is 7.8, and maximal product yield during a 30-min incubation occurs at 40 °C. Activity in frozen cell extracts remains constant for 5 days, then falls slowly to one-third of the initial value after 3 months. Enzyme activity rapidly declines irreversibly in the absence of pyridoxal phosphate. Agarose gel chromatography of the native enzyme yields a single band of activity at an elution volume corresponding to a molecular weight of 138,000. δ-Aminolevulinic acid synthase obtained from green wild-type strain Z cells is identical in its physical properties to that obtained from white aplastidic mutant strain W14 ZNalL cells. 相似文献
8.
Lifetable demography and reproductive traits of a Kenyan strain of the rotifer Brachionus angularis were investigated using individual and small batch culture approaches. The rotifer was identified morphologically before conducting studies at 20, 25 and 30 °C, using Chlorella vulgaris at 2.5 × 105 to 2.5 × 107 cells ml–1. The rotifers were highly fecund, producing 2.11 ± 0.07 offspring female–1 day–1 and reproductive, producing 8.43 ± 0.24 offspring female–1 at 25 °C with 2.5 × 106 algal cells ml–1. The highest intrinsic rate of natural increase (0.74 ± 0.02 d–1), specific population growth rate (0.49 ± 0.01), longest life expectancy at hatching (12.41 ± 0.28 d) and shortest generation time (2.87 ± 0.03 d) also occurred at 25 °C with 2.5 × 106 algal cells ml–1. The duration of hatching to first spawning was shortest (2.86 ± 0.21 h) at 30 °C with 2.5 × 107 algal cells ml–1 and longest (8.83 ± 0.39 h) at 20 °C with 2.5 × 105 algal cells ml–1. The highest population density (255.7 ± 12.6 ind. ml–1) was realised at 25 °C with 2.5 × 106 cells ml–1 on Day 8, whereas the lowest population density (122.0 ± 3.6 ind. ml–1) was realised at 20 °C with 2.5 × 105 cells ml–1 on Day 8. The lorica length and width of the Kenyan strain of B. angularis are 85.6 ± 3.1 µm and 75.4 ± 3.6 µm, respectively. The rotifer optimally reproduces at 25 °C when fed with 2.5 × 106 algal cells ml–1. 相似文献
9.
Young-Kwon Seo Doo-Hoon Lee Youn-Ho Shin Bo-Young You Kyung-Mi Lee Key-Yong Song Seong-Jun Seo Sung-Joo Whang Young-Jin Kim Eun-Kyung Yang Chang-Seo Park Ih-Seop Chang Jung-Keug Park 《Biotechnology and Bioprocess Engineering》2003,8(2):151-157
Obtaining a sufficient amount of healthy keratinocytes from a small tissue is difficult. However, ORS cells can be a good
source of epithelium since they are easily obtainable and patients do not have to suffer from scar formation at donor sites.
Accordingly, the current study modified the conventional primary culture technique to overcome the low propagation and easy
aging of epithelial cells during culturing. In a conventional primary culture, the average yield of human ORS cells is 2.1×103 cells/follicle based on direct incubation in a trypsin (0.1%)/EDTA (0.02%) solution for 15 min at 37°C, however, our modified
method was able to obtain about 6.9×103 cells/follicle using a two-step enzyme digestion method involving dispase (1.2 U/mL) and a trypsin (0.1%)/EDTA (0.02%) solution.
Thus, the yield of primary cultured ORS cells could be increasd three times higher. Furthermore, a total of 2.0×107 cells was obtained in a serum-free medium, while a modified E-medium with mitomycin C-treated feeder cells produced a total
of 6.3×107 cells over 17 days when starting with 7.5×104 cells. Finally, we confirmed the effectiveness of our ORS cell isolation method by presenting their ability for reconstructing
the bioartificial skin epitheliumin vitro 相似文献
10.
Allison A. Welder Roberta Grant June Bradlaw Daniel Acosta 《In vitro cellular & developmental biology. Animal》1991,27(12):921-926
Summary Tricyclic antidepressants (TCAs) are currently used in the treatment of mental depression and nocturnal enuresis. Clinically,
these drugs are useful; however, cardiotoxicity can occur even with therapeutic dosages. For example, TCAs are known to alter
myocardial function, induce arrhythmias, and produce heart block in individuals with a normal cardiovascular history. The
present study was undertaken to establish a culture system of spontaneously contracting adult primary myocardial cells for
toxicologic testing and to examine their contractility, morphology, and lactate dehydrogenase release (LDH) after treatment
with one of the most cardiotoxic TCAs, amitriptyline. Primary myocardial cell cultures were obtained from approximately 60-
to 90-day-old Sprague-Dawley rats. After the cells had been grown in culture for 11 days, they were treated with amitriptyline
(1 × 10−3, 1 × 10−4, and 1 × 10−5
M) for 2 to 24 h. The highest concentration of amitriptyline (1 × 10−3
M) completely destroyed the cardiac muscle cells. In addition to moderate and severe vacuole, granule, and pseudopodia formation,
all contractile activity was inhibited as early as 2 h after exposure to the intermediate concentration of 1 × 10−4
M amitriptyline. Significant LDH release did not occur until 8 h after treatment with this intermediate concentration. Even
though there was no significant LDH release at all 3 time points tested, there was a 50% decrease in beating activity (154±9
to 77±5 beats/min) and initiation of vacuole formation by 2 h with the lowest concentration of amitriptyline (1 × 10−5
M). This study presents a new apparatus for the isolation of adult cardiac myocytes for the establishment of primary cell cultures
for toxicologic testing. Furthermore, these data demonstrate that amitriptyline induces a concentration- and time-dependent
cardiotoxic profile in a model of spontaneously contracting adult cardiac muscle cells in culture. 相似文献
11.
Amanda Mizukami Maristela D. Orellana Sâmia R. Caruso Karen de Lima Prata Dimas T. Covas Kamilla Swiech 《Biotechnology progress》2013,29(2):568-572
The need for efficient and reliable technologies for clinical‐scale expansion of mesenchymal stromal cells (MSC) has led to the use of disposable bioreactors and culture systems. Here, we evaluate the expansion of cord blood‐derived MSC in a disposable fixed bed culture system. Starting from an initial cell density of 6.0 × 107 cells, after 7 days of culture, it was possible to produce of 4.2(±0.8) × 108 cells, which represents a fold increase of 7.0 (±1.4). After enzymatic retrieval from Fibra‐Cell disks, the cells were able to maintain their potential for differentiation into adipocytes and osteocytes and were positive for many markers common to MSC (CD73, CD90, and CD105). The results obtained in this study demonstrate that MSC can be efficiently expanded in the culture system. This novel approach presents several advantages over the current expansion systems, based on culture flasks or microcarrier‐based spinner flasks and represents a key element for MSC cellular therapy according to GMP compliant clinical‐scale production system. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 29: 568–572, 2013 相似文献
12.
Regeneration of Mesophyll Protoplasts Isolated from Dihaploid Clones of Solanum tuberosum 总被引:3,自引:0,他引:3
Protoplasts have been isolated from leaves of shoot cultures of six dihaploid clones of Solanum tuberosum L. (2n = 2x = 24). In the KM medium (Kao and Michayluk 1975), sustained cell divisions were obtained in up to 50% of the plated protoplasts of four clones, whereas only a few divisions occurred in the other two clones. The first mitosis appeared 2–8 days after plating, dependent on the clones. In the clones showing sustained cell divisions, a protoplast titre of about 5 × 103 per ml turned out to be optimal. The culture conditions for protoplasts of one of the poorly growing clones, clone H2 140, have been improved using modified KM media, plating at a concentration of as high as 5 × 104 cells per ml, and subsequent diluting at intervals 5 days. The dilutions were carried out with media containing 0.25% agar. Up to 60% of the plated protoplasts underwent divisions within 10 days under these conditions. After about 15 days, the regenerants were transferred onto media inducing organogenesis. Shoots and roots were formed on modified media MS (Murashige and Skoog 1962) and B5 (Gamborg et al. 1968). Plants have been regenerated in four of the investigated clones. Countings of chromosomes revealed a satisfactory stability of the karyotype in shoot culture and protoplast regeneration. 相似文献
13.
In this paper, we describe a method for primary culture of a well differentiated electrically tight rabbit vocal fold epithelial cell multilayer and the measurement of transepithelial electrical resistance (TEER) for the evaluation of epithelial barrier function in vitro. Rabbit larynges were harvested and enzymatically treated to isolate vocal fold epithelial cells and to establish primary culture. Vocal fold epithelial cells were co-cultured with mitomycin C-treated feeder cells on collagen-coated plates. After 10–14 days in primary culture, cells were passaged and cultured until they achieved 70–90% confluence on collagen-coated plates. Epithelial cells were then passaged onto collagen-coated cell culture inserts using 4.5 cm2 membrane filters (1.0 μm pore size) with 10% fetal bovine serum or 30 μg/mL bovine pituitary extract to investigate the effects of growth-promoting additives on TEER. Additional experiments were performed to investigate optimal seeding density (1.1, 2.2, 4.4, or 8.9 × 105 cells/cm2), the effect of co-culture with feeder cells, and the effect of passage number on epithelial barrier function. Characterization of in vitro cultures was performed using hematoxylin and eosin staining and immunostaining for vocal fold epithelial cell markers and tight junctions. Results revealed higher TEER in cells supplemented with fetal bovine serum compared to bovine pituitary extract. TEER was highest in cells passaged at a seeding density of 2.2 × 104 cells/cm2, and TEER was higher in cells at passage two than passage three. Ultrastructural experiments revealed a well-differentiated epithelial cell multilayer, expressing the epithelial cell markers CK13, CK14 and the tight junction proteins occludin and ZO-1. 相似文献
14.
B. Shi S. Zhang Y. Wang Y. Zhuang J. Chu S. Zhang X. Shi J. Bi M. Guo 《Cell proliferation》2010,43(3):275-286
Background: Sertoli cells (SCs) have been described as the ‘nurse cells’ of the testis whose primary function is to provide essential growth factors and create an appropriate environment for development of other cells [for example, germinal and nerve stem cells (NSCs), used here]. However, the greatest challenge at present is that it is difficult to obtain sufficient SCs of normal physiological function for cell transplantation and biological medicine, largely due to traditional static culture parameter difficult to be monitored and scaled up. Objective: Operational stirred culture conditions for in vitro expansion and differentiation of SCs need to be optimized for large‐scale culture. Materials and methods: In this study, the culturing process for primary SC expansion and maintaining lack of differentiation was optimized for the first time, by using microcarrier bead technology in spinner flask culture. Effects of various feeding/refreshing regimes, stirring speeds, seed inoculum levels of SCs, and concentrations of microcarrier used for expansion of mouse SCs were also explored. In addition, pH, osmotic pressure and metabolic variables including consumption rates of glucose, glutamine, amino acids, and formation rates of lactic acid and ammonia, were investigated in culture. Results: After 6 days, maximal cell densities achieved were 4.6 × 106 cells/ml for Cytodex‐1 in DMEM/FBS compared to 4.8 × 105 cells/ml in static culture. Improved expansion was achieved using an inoculum of 1 × 105 cells/ml and microcarrier concentration of 3 mg/ml at stirring speed of 30 rpm. Results indicated that medium replacement (50% changed everyday) resulted in supply of nutrients and removal of waste products inhibiting cell growth, that lead to maintenance of cultures in steady state for several days. These conditions favoured preservation of SCs in the undifferentiated state and significantly increased their physiological activity and trophic function, which were assessed by co‐culturing with NSCs and immunostaining. Conclusion: Data obtained in this study demonstrate the vast potential of this stirred culture system for efficient, reproducible and cost‐effective expansion of SCs in vitro. The system has advantages over static culture, which has major obstacles such as lower cell density, is time‐consuming and susceptible to contamination. 相似文献
15.
An ice-nucleating bacterium, strain KUIN-1, was isolated from the leaves of field beans (Phaseolus vulgaris L.). Strain KUIN-1 was identified as Pseudomonas fluorescens from its taxonomical characteristics. Ice-nucleating activity was obtained when strain KUIN-1 was cultured aerobically in a medium containing Koser citrate broth (pH 7.0) for 24 hr at 18°C. The ice- nucleating activity did not appear until the bacterial cell concentration reached 107 to 108/ml. Nucleation at — 3.0°C was detected in suspensions (1.8 × 109 cells/ml) of cells that had been grown on the medium containing Koser citrate broth. Strain KUIN-1 produced a lower nucleation frequency (i.e. the number of ice nuclei/cell) than did ice-nucleating Pseudomonas syringae No. 31 suspensions, particularly at temperatures above — 5°C. The nucleation frequency of strain KUIN- 1-suspensions was similar to that obtained for an ice-nucleating Erwinia herbicola No. 26 at — 5°C. 相似文献
16.
Gabriel Moura Mascarin Sérgio Batista Alves Fátima Teresinha Rampelotti-Ferreira Mariana Ragassi Urbano Clarice Garcia Borges Demétrio Italo DelaliberaJr. 《BioControl》2010,55(5):657-671
In this study a Brazilian granulovirus strain, PhopGV, isolated from the potato tuber moth (PTM) Phthorimaea operculella, was investigated regarding its potential for biological control and in vivo production. The relationship between mortality
of P. operculella larvae and virus concentration was determined at different temperatures on potato tubers and susceptibility of P. operculella to PhopGV was also determined on potato leaves. Virulence of PhopGV to P. operculella was not affected by temperatures from 18 to 30°C. The median lethal concentration (LC50) of larvae fed on potato foliage treated with PhopGV was not higher than that verified with larvae fed on treated tubers.
Optimal conditions for production of virus-infected larvae were obtained by using the virus suspensions of 41 × 105, 6.3 × 105 and 62 × 105 OBs ml−1 at 18, 24 and 30°C, which resulted in 32.0, 31.4 and 34.8% of infected larvae collected, respectively. The maximum percentage
of infected larvae recovered from tubers was not affected by temperature. However, time for production of virus-infected larvae
was longer at 18°C and shorter at 30°C. Persistence of PhopGV was determined on stored tubers and we observed that the virus
remained effective for at least two months, causing up to 84.2% mortality of P. operculella at 1 × 107 OBs ml−1. The pathogen was also highly virulent to tomato pinworm, Tuta absoluta, inflicting high percentage of mortality, delaying larval growth and inhibiting pupation. This Brazilian PhopGV strain has
potential to control PTM larvae on potato tubers at a broad range of temperature and can be produced in vivo using virus-treated
tubers. 相似文献
17.
Fish keratocytes are used as a model system for the study of the mechanics of cell motility because of their characteristic rapid, smooth gliding motion, but little work has been done on the regulation of fish keratocyte movement. As TGFβ (transforming growth factor β) plays multiple roles in primary human keratinocyte cell migration, we investigated the possible involvement of TGFβ in fish keratocyte migration. Studying the involvement of TGFβ1 in 24 h keratocyte explant allows the examination of the cells before alterations in cellular physiology occur due to extended culture times. During this initial period, TGFβ levels increase 6.2‐fold in SFM (serum‐free medium) and 2.4‐fold in SFM+2% FBS (fetal bovine serum), while TGFβ1 and TGFβRII (TGFβ receptor II) mRNA levels increase ~3‐ and ~5‐fold respectively in each culture condition. Two measures of motility, cell sheet area and migration distance, vary with the amount of exogenous TGFβ1 and culture media. The addition of 100 ng/ml exogenous TGFβ1 in SFM increases both measures [3.3‐fold (P=4.5 × 10?5) and 26% (P=2.1 × 10?2) respectively]. In contrast, 100 ng/ml of exogenous TGFβ1 in medium containing 2% FBS decreases migration distance by 2.1‐fold (P=1.7 × 10?7), but does not affect sheet area. TGFβ1 (10 ng/ml) has little effect on cell sheet area in SFM cultures, but leads to a 1.8‐fold increase (P=1.5 × 10?2) with 2% FBS. The variable response to TGFβ1 may be, at least in part, explained by the effect of 2% FBS on cell morphology, mode of motility and expression of endogenous TGFβ1 and TGFβRII. Together, these results suggest that expression of TGFβ and its receptor are up‐regulated during zebrafish keratocyte explant culture and that TGFβ promotes fish keratocyte migration. 相似文献
18.
Tiago B. Ferreira Ricardo Perdigão Ana C. Silva Changhe Zhang John G. Aunins Manuel J.T. Carrondo Paula M. Alves 《Biotechnology progress》2009,25(1):235-243
As the market requirements for adenovirus vectors (AdV) increase, the maximisation of the virus titer per culture volume per unit time is a key requirement. However, despite the fact that 293 cells can grow up to 8 × 106 cell/mL in simple batch mode operations, for optimal AdV infection a maximum cell density of 1 × 106 cell/mL at infection time has usually been utilized due to the so called “cell density effect”. In addition, AdV titer appears to be dependent upon cell cycle phase at the time of infection. To evaluate the dependence of AdV production upon cell cycle phase, 293 cells were chemically synchronised at each phase of the cell cycle; a 2.6‐fold increase on AdV cell specific titer was obtained when the percentage of cells at the S phase of the cell cycle was increased from 36 to 47%; a mathematical equation was used to relate AdV cell specific productivities with cell synchronisation at the S phase using this data. To avoid the use of chemical inhibitors, a temperature shift strategy was also used for synchronisation at the S phase. S phase synchronisation was obtained by decreasing the culture temperature to 31°C during 67 h and restoring it to 37°C during 72 h. By using this strategy we were able to synchronise 57% of the population in the S phase of the cell cycle obtaining an increase of 7.3‐fold on AdV cell specific titer after infection. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009 相似文献
19.
A. Meijer C. E. Vallinga J. M. Ossewaarde 《Applied microbiology and biotechnology》1996,46(2):132-137
We studied the propagation of Chlamydia pneumoniae strain TW-183 in HEp2 cells grown on microcarrier beads. Infection of the cells in microcarrier culture was optimized by
addition of 7.5% polyethylene glycol 4000 (PEG4000) during adsorption. The yield in microcarrier culture was similar to that
of microtitre-plate culture using centrifugation-assisted infection (120×106 and 225×106 bacteria/106 HEp2 cells respectively), as was the burst size (505 and 449 bacteria produced/infecting bacterium respectively). However,
up to 64% savings in labour time and 27% savings in culture medium were achieved if the microcarrier culture method was used
instead of the microtitre-plate culture method. The optimal yield of viable bacteria could only be achieved at a narrow range
of multiplicities of infection (0.24–1.14 inclusion-forming units/cell), independent of the mode of infection (centrifugation-assisted
infection or PEG4000-facilitated infection by adsorption) and independent of incubation temperature (35°C or 37°C). The yield
of microcarrier cultures was the same at an incubation temperature of 35°C or 37°C in contrast to an increased production
at 35°C in the microtitre-plate culture method using centrifugation-assisted infection. In conclusion, the microcarrier culture
method is useful to produce large quantities of viable Chlamydia pneumoniae economically.
Received: 27 December 1995/Received revision: 4 April 1996/Accepted: 15 April 1996 相似文献
20.
S H Golub 《Cellular immunology》1977,28(2):371-378
Culture conditions have been established that result in the sensitization of normal human peripheral blood lymphoid cells on allogeneic melanoma monolayers. Optimal culture conditions require 2 to 8 × 106 mitomycin C treated stimulator melanoma cells to sensitize 5 to 10 × 106 responder lymphoid cells. Neither rocking nor refeeding of the culture is necessary for the sensitization procedure. Stimulator cells grown in either fetal calf serum or human serum will serve as effective stimulator or target cells. Peak cytotoxic activity was detected at 44 hr in a microcytotoxicity assay, although some cytotoxic activity was detectable at 24 hr. 相似文献