SEARCH · PubMed Health

Results for “Serial Passage”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 829 records · Page 46Linked to original sources

PubMed1997

Characterization of Shigella type 1 fimbriae: expression, FimA sequence, and phase variation.

This study documents the presence of type 1 fimbriae on Shigella and confirms these mannose-sensitive adherence structures to be bona fide components of the Shigella surface. While laboratory-passaged Shigella strains and lyophilized clinical isolates failed to express type 1 fimbriae, 6 of 20 recent clinical isolates, including 4 Shigella flexneri strains, 1 Shigella boydii strain, and 1 Shigella dysenteriae strain, produced type 1 fimbriae as detected by mannose-sensitive hemagglutination (MSHA) and electron microscopy. Optimal production of a predominantly Fim+ population required serial passage every 48 to 72 h in unshaken brain heart infusion broth at 37 degrees C. Fim+ Shigella cultures were capable of reversibly switching to a non-MSHA, afimbriated phase during serial aerobic cultivation on tryptic soy agar plates. The amino acid sequence of S. flexneri type 1 FimA contained 18 substitutions compared to that of Escherichia coli fimbrillin. Indirect immunoelectron microscopy suggested the presence of both shared and unique epitopes on E. coli and S. flexneri type 1 fimbriae. Random phase variation between fimbriated and afimbriated states in Shigella was accompanied by the genomic rearrangement associated with phase variation in E. coli.

Amino Acid Sequence↗

PubMed1998

Random population-wide genetic damage induced in replicating cells treated with methotrexate.

Low lethality treatment of the NIH 3T3 mouse cell line with methotrexate (MTX) during exponential multiplication results in heterogeneous, heritable reduction in growth rate of most if not all the replicatively surviving cells. The effective concentrations of MTX are 10 to 100 times higher in molecular, cellular and developmental biology medium 402 (MCDB 402) than in Dulbecco's modification of Eagle's medium (DMEM) medium because of the folate-sparing presence of adenine, thymidine and, particularly, of folinic acid in MCDB 402 medium. The reduced growth rates are detectable during early passages of surviving populations before the faster growing cells dominate them. The heritable effect is most clearly demonstrated by sequestered cloning of many individual cells immediately after drug treatment, and repeatedly measuring the growth rates of the clones in serial passages. After 7-10 passages of the clones, there is an increase in growth rate of some of the slow growing clones presumably due to the generation and selection of faster growing cells. Evidence from mutagenic studies at a single genetic locus in other cell lines suggests that heritable reductions in growth rate arise from chromosome aberrations although point mutations may also contribute to the effect. Clastogenic changes can be induced by a wide variety of mutagens and carcinogens, many of which are used in chemotherapy of cancer and other chronic diseases. The population-wide, heritable damage to cells may be the source of, or may contribute to, late-occurring side effects of treatment in cancer and other chronic diseases.

3T3 Cells↗

PubMed1969

Propagation of MM virus in continuous cell lines.

Baby hamster kidney (BHK), McCoy, and L cell lines were found to be suitable for isolation of MM virus from infected mouse brain tissue. The virus was recovered in high titer in the first passage in BHK and McCoy cells, with concomitant cytopathic effect (CPE). In L cells, virus yield was lower than in the other two cell lines and CPE was incomplete. Adaptation of the virus to BHK and McCoy cells by serial passages was evidenced by accelerated development of the CPE and increase in the virus titer. Plaques were obtained in all three cell lines when inoculated with infected mouse brain or with the tissue culture-propagated virus. In the BHK cells, the virus release preceded the appearance of CPE and maximal yield of virus was obtained after 1 to 3 days of incubation, depending on the size of inoculum. The BHK-propagated virus had the same lethality for mice as did the mouse brain-propagated stock, and there was no difference in the course of the disease caused by the two preparations.

Adaptation, Biological↗

PubMed2004

Elevated telomerase activity and minimal telomere loss in cord blood long-term cultures with extensive stem cell replication.

Telomerase activity, telomere length, stem/progenitor cell production, and function of CD34+ cells from cord blood (CB), bone marrow, and mobilized peripheral blood were evaluated in long-term cultures. CB cells were cultured either on OP-9 stromal cells transduced with an adenovector expressing thrombopoietin (TPO) or stimulated by a cytokine cocktail in the absence of stroma, with, in one method, CD34+ cells reisolated at monthly intervals for passage. Continuous expansion of stem cells as measured by in vitro cobblestone area and secondary colony-forming assays was noted for 18 to 20 weeks and by severe combined immunodeficiency (SCID)-repopulating cells (SRCs), capable of repopulating and serially passage in nonobese diabetic/SCID mice, for 16 weeks. Despite this extensive proliferation, telomere length initially increased and only at late stages of culture was evidence of telomere shortening noted. This telomere stabilization correlated with maintenance of high levels of telomerase activity in the CD34+ cell population for prolonged periods of culture. Cytokine-stimulated cultures of adult CD34+ cells showed CD34+ and SRC expansion (6-fold) for only 3 to 4 weeks with telomere shortening and low levels of telomerase. There is clearly a clinical value for a system that provides extensive stem cell expansion without concomitant telomere erosion.

Adenoviridae↗

PubMedDate not supplied

[A mechanism for limiting reproduction influenza A virus reassortants with incomplete functional compatibility of hemagglutinin and neuraminidase gene products].

The mechanism of decrease in the level of virus accumulation in reassortants with hemagglutinin (HA) and neuraminidase (NA) genes from different parents is studied. The reassortant viruses and their passage variants do not differ by the rate of virus protein production or their stability in infected cells. Electron microscopy and titration of infectious virus in culture fluid and cell-associated virus showed that the variants selected by serial passages accumulated mainly in the culture fluid, whereas the initial reassortant virions were predominantly cell-associated. These data suggest that incomplete removal of sialic acid residues by viral neuraminidase N1 in some reassortants results in re-attachment of virions to the infected cells and thus impairs the virus dissemination, which may be regarded as a reassortant-limiting factor probably significant for virus evolution.

Animals↗

PubMed1987

Restriction endonuclease analysis of herpes simplex virus from recrudescent lesions, from latent infection and during passage in the skin and nervous system of mice.

The stability of the restriction endonuclease profile of herpes simplex virus type 1 strain SC16 in mice was studied. Virus isolated from skin during acute infection was compared with that from latently infected ganglia and with that from recrudescent lesions induced by trauma. In another experiment virus serially passaged only in skin was compared with virus that had also replicated in the nervous system. The loss or gain of specific restriction sites was not observed but in some cases the mobility of certain fragments decreased.

Animals↗

PubMed1962

Live tissue-culture influenza vaccine.

With a view to improving the quality of live influenza vaccines (previously passaged on eggs) in the USSR, attempts were made to obtain vaccine strains of virus by recent tissue-culture methods.Strains of A2 and B viruses obtained from patients in influenza epidemics were isolated on chick embryos and cultured by serial passage on monkey-kidney tissue or skin-muscle tissue of human embryos; a variant strain was obtained by further passaging of tissue-culture virus fluid through chick embryos. These strains all became gradually adapted to the tissue-culture and were generally stabilized by the 10th to 13th passage.Intranasal insufflation of the vaccine into volunteers resulted in twofold to eightfold antibody increases in the great majority and gave rise to almost no reactions.Lyophilization of the strains with sucrose-gelatin or calf serum ensured the stability of the vaccines for periods of up to six months.

Administration, Intranasal↗

PubMed1987

The effect of cell passage on the susceptibility of BALB/3T3 clone A31-1-1 cells to 3-methylcholanthrene-induced morphological transformation.

The response of BALB/3T3 clone A31-1-1 cells to chemically induced morphological transformation was evaluated using 3-methylcholanthrene (MCA). Stock cultures were initiated from cryopreserved cells, grown in T25 flasks containing 5 ml of medium, and replated at subconfluency. Serially transferred cells were then subjected to transformation assay. After 24-hr seeding, cells were incubated 48 hr with MCA in a 5% CO2 incubator. They were then rinsed and incubated for an additional 4 weeks with twice weekly medium change. Type III foci were scored after fixation and staining with Giemsa. With serial passage from the frozen state, cells of passages 3-14 had a low level of spontaneous transformation; zero to 6 type III foci per 20 dishes were counted. In the MCA-treated cultures the number of transformed foci, however, increased with passage. Such passage-related sensitivity to MCA was demonstrated for cells cultured in two batches of sera: one from MA Bioproducts (Lot no. 2E052) and the other from Armour Pharmaceuticals (Lot no. Y65801). The passage-related increase in number of transformed foci was not related to doubling time, cloning efficiency, or MCA-induced growth inhibition.

Animals↗

PubMed1982

Interactions of insulin with bovine endothelium.

Bovine endothelial cells have been isolated from pulmonary and systemic vessels and grown in culture as primary, passaged and passaged cloned-strains. The cultures were shown to be endothelial in nature on the basis of several endothelial-specific and endothelial-associated traits. Endothelial cells from all sources had specific receptors for insulin in primary culture and after serial passage. Endothelial cells derived from pulmonary arteries and aortas bound 2.5 times more insulin than cells derived from the pulmonary vein. Each endothelial cell type maintained a specific complement of receptors through at least 25 passages in vitro. Coupled with previous findings of insulin receptors on endothelial cells from human umbilical vessels, these data suggest that insulin receptors may be an intrinsic component of all vascular endothelium.

Animals↗

PubMed1997

Tissue culture adaptation of natural isolates of simian virus 40: changes occur in viral regulatory region but not in carboxy-terminal domain of large T-antigen.

The regulatory region of natural isolates of simian virus 40 (SV40) is different from that of laboratory-adapted strains of the virus. The latter have a nucleotide sequence duplication within the enhancer region which varies slightly with each strain, whereas the duplication is lacking in fresh isolates of SV40, which contain an 'archetypal' regulatory region. Many isolates also display nucleotide differences in the DNA encoding the carboxy terminus of large tumour antigen (T-ag). To determine whether genetic changes in these two regions of the SV40 genome were detectable during laboratory adaptation and long-term passage, low-passage virus stocks of two laboratory strains which had detailed passage histories spanning more than 25 years (Baylor strain and VA45-54) were analysed using PCR, cloning and sequencing assays. Both laboratory and archetypal regulatory regions were present in low-passage stocks. Following duplication in the regulatory region, no additional changes were detectable. The variable region at the T-ag carboxy terminus did not undergo any change with tissue culture passage and may serve as a useful site for taxonomic classification of different strains of SV40. Cloned genomes containing single or duplicated enhancers derived from both SV40 strains were viable in CV-1 cells. Attempts to induce regulatory region duplications by 14 serial passages of SV40 archetypal strains in monkey cells were not successful. The results are compatible with tissue culture adaptation of SV40, reflecting either selection of a rare variant pre-existing in the original sample or generation of a rare regulatory region duplication in infected cells.

Adaptation, Physiological↗

PubMed1984

Repeated passage of freshly isolated group A streptococci on blood agar. I. Effect on M protein, opacity factor and IgG Fc-receptor activity.

Seventy-six strains, 20 T-type 1, 20 T-type 2, 18 T-type 4 and 18 T-type 12 group A streptococci were isolated from throat cultures and subjected to 25 serial passages on blood agar. A single "glossy" colony was selected from each passage in order to diminish the M protein content of the strains. In accordance with previous results, the M protein synthesis, as estimated by an electro-immuno assay diminished in the T1 strains, in mean from 23.8 to 15.5% of a reference M protein preparation (p less than 0.01). Opacity factor (OF)-production was used as a measure for M protein in the T-type 2, the T-type 4 and the OF-positive T-type 12 strains. OF-excretion decreased significantly in the T-types 2 and decreased, though not significantly in the T-type 12 strains but increased (unexpectedly) in the T-type 4 isolates during subculture. However, irrespective of the changes in M protein/OF production these types all increased significantly in capacity to bind radiolabelled IgG via the Fc-fragment. It is known that streptococci isolated during convalescence resemble subcultured strains; it is suggested that IgG Fc-receptors are important for protection of the streptococci against phagocytosis in the asymptomatic carrier state.

Humans↗

PubMed1976

Endotoxin-induced non-specific resistance in rats examined by Trypanosoma equiperdum challenge.

In rats weighing 100 g, intravenously administered Trypanosoma equiperdum organisms started logarithmic growth immediately, whereas in rats weighing 150-300 g growth started after a lag phase of 0.7-3.1 hr. The lag phase lasted for 15-17 hr when the rats were pretreated serially with endotoxins and this time course was not modified by changes either in the host's body weight or in the germ count of the inocula. Maximum resistance to infection was achieved with gradual doses of the endotoxin given at 48 hr intervals on 6 occasions. The total dose of endotoxin was fourfold of its LD50. Additional doses failed further to increase the resistance. Serial passages of the strain in rats pretreated with endotoxin disclosed that the trypanosomes growing after the lag phase had developed resistance to the factor responsible for the lag phase. Thus the method does not allow a quantitative estimation of resistance induced by serial endotoxin treatment, yet it represents a rapid and simple procedure in every case in which qualitative assessment is sufficient.

Animals↗

PubMed1976

Avian infectious bronchitis vaccine: primordial or derived virus?

Vaccine formulations of virulent, modified/attenuated, or inactivated mono- or multivalent types of chicken embryo-propagated avian infectious bronchitis virus (IBV) at different passage levels and administered by non-parenteral routes have been used for immunization of chickens against the disease. Two doses of vaccines are generally used: 1) high level virus of lessened virulence, antigenicity and immunogenicity as "derived" from its parental virus via serial passage in and selective adaptation to the chicken embryo, and 2) low passage level virus with the above properties more closely related to the greater potency of the "primordial" parental virus in nature. The efficacy of vaccines with contemporary or candidate virus has been interpreted from a variety of criteria among which are: 1) clinical response, residence and dissemination of virus, 2) in vivo and in vitro local tissue response of tracheal epithelium, 3) protection against virulent homologous or heterologous types of IBV, and 4) induced secretory and humoral antibody. Vaccines have been economically useful and of epizootiological value but the greatest difficulty is the uncertainty of protection because of the changing antigenic, immunogenic and other properties of existing virus and of newly emerging types. With a complex virus such as IBV, the opportunities for natural selection and evolution of new or modified antigens via association with antibody of immune or of partially immune hosts are as enormous as antigenic lability resulting from fragmentation of the primordial genome under natural and/or artificial environmental stress. The present day IBV might be progeny from a unique pool of primordial genome via mutation and host-induced variation, or even persistence of primordial virus as an infinitesimal portion of the population.

Animals↗

PubMed1962

Antibody-producing capacity of adult chicken spleen cells in newly hatched chicks.

1. To evaluate the effect of primary in vitro antigenic stimulation on a population of immunologically competent cells, a homologous cell transfer system was used, with adult chickens as the spleen cell donors, killed Brucella abortus as the antigen, and newly hatched chicks as the recipients. 2. The distribution of antibody titers in recipients of cells from random donors was bimodal, with about 30 per cent showing no detectable titer and the remainder distributed normally. 3. Variability of titer was reduced significantly in subgroups receiving cells from a single donor, indicating that the primary source of variability was the genetic capacity of discrete cell populations to respond to antigen. 4. In serial passage studies, activity of donor cell populations was lost rapidly: antibody was not demonstrated after the second passage, and the graft versus host reaction (splenomegaly) was not demonstrated after the third passage. Results were similar with in vitro antigenic stimulation at the time of the first passage only and with additional stimulation at the time of subsequent transfers. 5. The thesis that,the homograft reaction of the newly hatched recipients had contributed significantly to the variability in the single transfer studies and to the rapid loss of activity in the serial transfer experiments was confirmed by the results of transfers following alteration of the lymphoreticular system of the host by thorotrast administration, splenectomy, and treatment with 19-nortestosterone during embryogenesis. All three favored the survival and function of transferred cells, raising the average antibody titer and virtually eliminating the no-response category. The inhibition of homograft immunity was most pronounced in the 19-nortestosterone group.

Animals↗

PubMed1988

Stage-selective inhibition of rodent malaria by cyclosporine.

The relative susceptibility of different developmental stages of Plasmodium berghei to cyclosporine was investigated in vivo. Within 12 h of receiving a single 25-mg/kg (body weight) dose of cyclosporine, mice with patent P. berghei infections uniformly exhibited a rapid fall in asexual parasite stages. Initially, ring forms and mature schizonts disappeared. Subsequently, trophozoites disappeared between 21 and 24 h, whereas gametocytes persisted for 36 h. In contrast, when cyclosporine was administered to mice 1 day before inoculation (100 mg/kg) with P. berghei sporozoites and for 2 consecutive days after inoculation (25 mg/kg), infections developed normally. When mice with patent infections were placed on prolonged cyclosporine therapy (25 mg/kg per day), parasitemia initially disappeared but often recrudesced. Recrudescent parasites were frequently resistant to cyclosporine (Csr). The Csr phenotype remained stable after serial passage of parasites in mice and after transmission through Anopheles stephensi mosquitoes, in which the capacity to produce oocysts was reduced. When infections of untreated mice were initiated with equal numbers of Csr and cyclosporine-susceptible (Css) parasites and then carried through two serial cycles of mosquito-to-mouse transmission without cyclosporine treatment, the Csr phenotype was lost. The results indicate that cyclosporine selectively inhibits asexual blood stages of P. berghei and favors the emergence of Csr parasites with diminished infectivity for mosquitoes.

Animals↗

PubMed2004

Characterization of foot-and-mouth disease serotype asial viruses grown in the presence of polyclonal antisera in serology and nucleotide sequence analysis.

Foot-and-mouth disease viruses (FMDV) have a high rate of mutation and spontaneous mutants can be readily. isolated in the laboratory. In this study, plaque purified FMDV Asial vaccine strains (IND 63/72 and IND 491/97) were passaged in-vitro in Baby Hamster Kidney-21 cell monolayers in the presence of sub-neutralizing levels of antiviral polyclonal sera (APS), raised in guinea pigs against the purified and inactivated whole virus particles of IND 63/72, IND 491/97 and IND 13/01. After serial passages under selective immune pressure, the viruses starts growing in the presence of undiluted sera and showed certain characteristics like an increased resistance to neutralization by APS and reduction in plaque counts on titration in plaque assay. Cross-neutralization of these viruses with above-mentioned APS revealed selection of three complete and one partial polyclonal antibody resistant (PAR) viruses based on the 'r' value in micro neutralization test. Alterations were detected at several amino acid residues in the structural protein-coding P1 region. Many of the residues inferred to be positively selected sites in other serotypes of this virus were also prone to substitution under immune selection pressure in Asia1 virus. The present work extends the finding that selection exerted by host antibody also plays a major role in the rapid evolution of FMDV Asia1, as observed in other serotypes.

Amino Acid Sequence↗

PubMed1986

Expression of epidermal differentiation antigens in cultures of interspecific somatic hybrids (human keratinocytes X mouse fibroblasts 3T3-4E).

Interspecific somatic hybrids have been prepared by fusion of human epidermal cells with mouse fibroblasts 3T3-4E using PEG 4000. Expression of epidermal differentiation antigens (bullous pemphigoid antigens, BP, keratin subsets 55-57 k and 67 k), markers of basal and suprabasal cells, were studied by immunocytochemistry for 10 passages. These markers were detected in the hybrids early after fusion, indicating that cells from both compartments were able to fuse with 3T3-4E cells. However, the hybrids expressing high molecular weight keratins were no longer detected after 7 days in primary cultures and serial passages, whereas those expressing BP antigens and vimentin persisted. Low molecular weight keratins 52 K and 50 K were detected by SDS-PAGE at the second passage in precipitates formed between labeled hybrid lysates and total keratin rabbit antiserum. Karyotype analysis showed mainly murine chromosomes and a submetacentric human chromosome between the 6th and the 10th passage.

Animals↗

PubMed1990

Noncorrelative c-myc and ras oncogene expression in squamous cell carcinoma cells with tumorigenic potential.

The distribution of heterogeneous cell types within human tumors was examined, and the biological behavior of tumors and different tumor cell lines was evaluated following implantation into surrogate hosts. In situ hybridization and immunohistochemistry were used to examine the expression of oncogenes and localization of the squamous cell carcinoma cell surface-associated antigens. Increased levels of H-ras mRNA and p21 protein were present in six tumors, but enhanced c-myc mRNA expression was observed in just two tumors. The distribution of oncogene mRNA and SCC antigen-positive cells was not uniform throughout the tumor. Isolation of cells from the tumors was accomplished by cell culture, growth in soft agar, and growth in the nude mouse. One nontumorigenic immortalized cell line, SCC-83-01-82, isolated by passage through soft agar, was treated with 50 micrograms/ml of methyl methane sulfonate (MMS). These MMS-converted cells subsequently expressed a tumorigenic phenotype. In situ hybridization of the tumors that developed in nude mice revealed increased c-myc and H-ras mRNA expression. Serial passage of the MMS-converted tumors in vivo was accompanied by consistent enhanced c-myc expression. However, the levels of H-ras and keratin mRNA expression decreased with passage in vitro. Northern blot analysis of c-myc and H-ras mRNA levels from the original SCC cell line showed no change in expression following MMS treatment. The data suggest that SCC-83-01-82 is a premalignant cell line established from a mixed cell population in the tumor mass. It can be converted to a malignant phenotype by treatment with MMS, and the persistence of malignancy is under molecular control other than changes in the level of c-myc and ras gene expression.

Animals↗