Background: The method of immunoelectron microscopy has been found more than 20 years. It is widely applied to detect and identify some types of virus in medical waste samples.\r\n', u'Objectives: To identify antigen location of Rota virus in organelle of the Vero cell and primary monkey kidney cells after infecting and to study the interaction between the virus and host cells.\r\n', u'Subjects and methods: The study was conducted on Rota virus G1P8 (KH0118) isolated from patients with symptoms of acute diarrhea, primary monkey kidney cells collected from Macaca mulatta monkey and the Vero cell of WHO. \r\n', u'Results: Gold particles (10nm) coated protein A and polyclonal antibodies were used to interact directly with Rotavirus proteins \r\n', u'These gold particles with high electron density revealed the antigen location of the Rota virus in the lysosome, pouch and other compartments of the cytoplasm.\r\n', u'Newly assembled viral particles could be identified only after 18-20hours post-infection. It is also noteworthy that viral particles and empty capsides (virus like particles) were comprised into cytoplasmic vesicles associated with the endoplasmic reticulum (ER)-Golgi system.\r\n', u'Conclusion: In order to better understand the interaction mechanism of virus and host cells, the use of this method together with specific monoclonal antibodies for each protein component of viruses and cells is essential.\r\n', u'\r\n', u'\r\n', u'
Rotavirus ; Vero Cells ;

Background: As avian influenza A/H5N1 epidemic spreads rapidly, many corporations, vaccine manufacturers in cooperation with laboratories around the world has conducted research and developed H5N1 vaccine for both humans and poultry. Objective: Evaluate the adaptation and cloning of rgh5n1 vaccine reference strain to vero cells in order to produce master seed virus and working seed virus. Subject and Method: The reverse genetics derived A/H5N1 virus strain (rgH5N1) was studied for adaptation to Vero cells by serial passage. It has been shown that the rgH5N1 strain can be propagated in Vero cells and cause CPE with the highest virus titer 1010,9 PFU/ml at Vero passage 15. The rgH5N1 strain was cloned by using plaque purification method and passaged to obtain a high stable virus titer. Conclusion: The reverse genetics derived A/H5N1 virus strain was propagated in Vero cells. Master seed virus and working seed virus were obtained at passage 6.
rgh5n1 ; avian influenza ; vero cells

No abstract available.
Humans* ; Sperm Injections, Intracytoplasmic* ; Spermatozoa* ; Vero Cells*

Background: \r\n', u'Rotavirus is the major important cause of infectious infantile gastroenteritis and diarrhea, especially the children aged 6 months to 24 months. Some efforts made to provide safe water and cultivate the hygiene have not reduced the diarrhea caused by rotaviruses. Develope a vaccine agaisnt this virus is of high importance. A question about the production of this kind of vaccine targeted athow to make human rotaviruses strains adapt cultured cells.\r\n', u'Objectives\r\n', u'To determine the multiplicity of infection (MOI) of human rotaviruses strains in vero cell.\r\n', u'Subjects and method: \r\n', u'The multiplicity of infection of Human rotaviruses strains (G1 P8, G1P4, and G4P6) were selected and the CDC- Atlanta US supplied the vero cell.Use vero cells cultivate, temper the vero cell and, applying direct immunofluorescence to determine the multiplicity of G1P8, G1P4, C4P6.\r\n', u'Results:\r\n', u'The MOI of G1P8, G1P4 and C4P6 are 0.02 ffu/cell, 0.15ffu/cell, and 0.35 ffu/cell, respectively.\r\n', u'Conclusion:\r\n', u'Using the vero cell to multiple the rotaviruses is the effective part during the production of vaccines against rotaviruses. The MOI of human rotaviruses strains G1P8, G1P4 and G4P6 are in order:0,02 ffu/cell, 0,15ffu/cell, and 0,35% ffu/cell. \r\n', u'
Rotavirus ; Vero Cells ; Cell Culture Techniques ;

Background: Vero cell (ATCC) is from kidney of Blue Monkey in Africa. Because of its strong points such as non tumor form, non exotic virus infection, this cell strain is commonly used for vaccin development in the world. Objective: To determine the quality of vero cell supplying by Japan Polio-myelitis Research Center and WHO vero cell supplying by the Company for Vaccine and Biological Production No.1 in use for develop rota vaccine. Subjects and method:A study was conducted in 2 kinds of vero cell (one ATCC 134 generation supplying by Japan Polio-myelitis Research Center and another WHO ATTC 137 generation supplying by the Company for Vaccine and Biological Production No.1) using standard methods. Results and Conclusion: Both these ATCCs had no exotic agents in generation from 134 to 137. The vero working cell bank for vaccine development has been established by the POLYVAC by using standard methods, in accordance with the WHO regulations. The vero working cells established by POLYVAC had the same quality as that of Vabiotech cell bank. Rota virus strains multiplied well on WHO ATTC 137 generation and ATCC 134 generation supplying by Japan Polio-myelitis Research Center.
Vero Cells/ drug effects ; Rotavirus Vaccines ;

Chrysin is a 5,7-dihydroxyflavone and was recently shown to potently inhibit enterovirus 71 (EV71) by suppressing viral 3C protease (3Cpro) activity. In the current study, we investigated whether chrysin also shows antiviral activity against coxsackievirus B3 (CVB3), which belongs to the same genus (Enterovirus) as EV71, and assessed its ability to prevent the resulting acute pancreatitis and myocarditis. We found that chrysin showed antiviral activity against CVB3 at 10 muM, but exhibited mild cellular cytotoxicity at 50 muM, prompting us to synthesize derivatives of chrysin to increase the antiviral activity and reduce its cytotoxicity. Among four 4-substituted benzyl derivatives derived from C(5) benzyl-protected derivatives 7, 9-11 had significant antiviral activity and showed the most potent activity against CVB3 with low cytotoxicity in Vero cells. Intraperitoneal injection of CVB3 in BALB/c mice with 1x106 TCID50 (50% tissue culture infective dose) of CVB3 induced acute pancreatitis with ablation of acinar cells and increased serum CXCL1 levels, whereas the daily administration of 9 for 5 days significantly alleviated the pancreatic inflammation and reduced the elevation in serum CXCL1 levels. Collectively, we assessed the anti-CVB3 activities of chrysin and its derivatives, and found that among 4-substituted benzyl derivatives, 9 exhibited the highest activity against CVB3 in vivo, and protected mice from CVB3-induced pancreatic damage, simultaneously lowering serum CXCL1 levels.
Acinar Cells ; Animals ; Enterovirus ; Inflammation ; Injections, Intraperitoneal ; Mice ; Myocarditis ; Pancreatitis ; Vero Cells

OBJECTIVE: The purpose of the study was to determine the effects of co-culture with oviductal epithelial cells and Vero cells on mouse embryo. METHOD: For the control group, mouse embryos were cultured alone in Ham's F-10 with 10% FBS. Subcultured oviductal epithelial cell and Vero cell were cocultured in Ham's F-10 with 10% FBS with the mouse embryo and used as the treatment group. Development of mouse embryos were observed. Result: The development rate and hatching rate of embryos that cocultured with oviductal epithelial cell and Vero cell was significantly higher (p<0.05) than control group. When subcultured oviductal epithelial cells were co-cultured with mouse embryo, there was no significant difference in development rate and hatching rate among subculture step. When oviductal epithelial cells that have been frozen-thawed were co-cultured with mouse embryo, there was no significant difference in development rate and hatching rate among subculture step. No statistical significance was seen in the development rate and hatching rate between subcultured oviductal epithelial cells and frozen-thawed oviductal epithelial cells when cocultured with mouse embryo, Vero cells and frozen-thawed when cocultured with mouse embryo, and Vero cells and oviductal epithelial cells when cocultured with mouse embryo. CONCLUSION: Oviductal epithelial cells and Vero cell may have a stimulatory role in early mouse embryonal development compared to control in vitro. As well, there is no significant difference in development rate and hatching rate among subculture step, when early mouse embryo was cocultured with cells that subcultured and frozen-thawed.
Animals ; Coculture Techniques ; Embryonic Structures ; Epithelial Cells* ; Humans* ; Mice* ; Oviducts* ; Vero Cells*

Recovery from potentially lethal damage (PLDR) after irradiation was studied in plateau-phase culture of Vero cells in vitro. Unfed plateau-phase cells were irradiated with dose of 1 to 9 gy using Cs-137 irradiator. Cells then were incubated again and left in situ for 0, 1, 2, 3, 4, 5, 6 and 24 hours and then were trypsinized, explanted, and subcultured in fresh RPMI-1640 media containing 0.33% agar. Cell survival was measured by colony forming ability. An adequate number of heavily irradiated Vero cells were added as feeder cells to make the total cell number constant in every culture dish. As the postirradiation in situ incubation time increased, surviving fraction increased saturation level at 2 to 4 hours after in situ incubation. As the radiation dose increased, the rate of PLDR also increased. In analysis of cell survival curve fitted to the linear-quadratic model, the linear inactivation coefficient (alpha) decreased largely and reached nearly to zero but the quadratic inactivation coefficient (beta) increased minimally by increment of postirradiation in situ incubation time. So PLDR mainly affected the damage expressed as alpha. In the multitarget model, significant change was not obtained in D0 but in Dq. Therefore, shoulder region in cell survival curve was mainly affected by PLDR and terminal slope was not influenced at all. And dose-modifying factor by PLDR was relatively higher in shoulder region, that is, in low dose area below 3 gy.
Agar ; Cell Count ; Cell Survival ; Feeder Cells ; Shoulder ; Trypsin ; Vero Cells*

This study was performed to observe the influence of the feeder layer on the culture of the human uveal melanoma cell. Two types of melanoma cells which were obtained from the patients with choroidal melanoma were implanted into the culture flask. The melanoma cell was seeded onto the culture dish covered with conjunctival fibroblast, Vero cell, and culture dish which did not contain the feeder layer respectively. The growth pattern of the melanoma cell was observed with phase contrast microscopy upto 30 days after seeding. There was no definite difference in growth pattern between each group. These results may indicate that the feeder layer is not an essential factor in the culture of uveal melanoma cell.
Choroid ; Feeder Cells ; Fibroblasts ; Humans ; Melanoma* ; Microscopy, Phase-Contrast ; Vero Cells

In order to establish an infectious clone for CDV-3, a commercial vaccine strain of canine distemper virus for mink, to provide reference for the studies of pathogenesis and novel vaccine development of CDV. Thirteen pairs of primers were used to amplify the full-length genome of CDV-3 strain. Five long fragments were obtained based on single restriction site analysis of the whole genome of CDV-3 by RT-PCR. Five fragments were successively inserted into the multiple clone sites in the modified eukaryotic vector of pcDNA3.2 by restriction enzymes and splicing. Meanwhile, the hammerhead ribozyme and hepatitis delta virus ribozyme sequences were added to the beginning of F1 fragment and the ending of F5 fragment, respectively. Then, the full-length cDNA recombinant plasmid of CDV-3 was obtained and named as pcDNA3.2-CDV-3. In addition, three helper plasmids, expressing the N protein, P protein and L protein of the CDV-3 strain respectively, were constructed. The 293T cells were transfected with the full-length cDNA recombinant plasmid and three helper plasmids by Lipofectamine™ 2000. At 3 days post transfection, the supernatant was added to the monolayer of Vero cells to observe the typical syncytium of CDV. Indirect immunofluorescence and artificial label identification of recombinant virus rCDV-3 were conducted after the occurrence of lesions. Finally, the growth characteristics of wtCDV-3 and rCDV-3 were compared after passaging of rCDV-3. The identification of the full-length cDNA recombinant plasmid and three helper plasmids by restriction enzyme digestion and sequencing were consistent with expected. The Vero cells infected with the recombinant rCDV-3 showed typical syncytic. The identification of indirect immunofluorescence and labeled marker, and observation under electron microscope proved that the rCDV-3 was indeed rescued from the recombinant plasmid of pcDNA3.2-CDV-3. In comparison of the virus titers of wtCDV-3, rCDV-3 replicated massively and rapidly and reached the maximize virus titer of 10⁷·⁶⁶⁷ TCID₅₀/mL within 36 h post infection (p.i.) in Vero cells, while wtCDV-3 grew gradually to 10⁶·⁶⁶⁷ TCID₅₀/mL at 72 h p.i. in Vero cells. This reverse genetic system of CDV-3 strain has been established successfully, to provide reference for the studies of pathogenesis and novel vaccine development of CDV.
Animals ; Chlorocebus aethiops ; Clone Cells ; DNA, Complementary ; Distemper Virus, Canine/genetics* ; Plasmids/genetics* ; Vero Cells