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Method for separating and culturing RSV (Respiratory Syncytial Virus) by using OMC (Ostiomeatal Complex)-685-1 cells

A syncytial virus, separation and culture technology, applied in the field of respiratory syncytial virus separation and culture, can solve the problems of low sensitivity, time-consuming, shortened culture time, etc., and achieve the effects of high sensitivity, improved sensitivity, and shortened time

Inactive Publication Date: 2012-07-11
ZUNYI MEDICAL UNIVERSITY
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Problems solved by technology

[0005] The technical problem to be solved in the present invention: solve the problems such as low sensitivity, time-consuming and long time-consuming of current RSV pathogen isolation, the present invention uses human ovarian mucinous cystadenocarcinoma cells ( OMC685-1) as a tool to isolate RSV virus, which improves the sensitivity of detection and shortens the incubation time compared with traditional virus isolation methods

Method used

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Embodiment 1

[0020] A method for separating and cultivating respiratory syncytial virus with OMC685-1 cells of the present invention comprises the following steps:

[0021] (1) OMC685-1 cell culture:

[0022] After recovering, OMC685-1 cells were inoculated in RPMI-1640 medium containing 10% calf serum, and placed in a relative humidity of 95﹪~98﹪, 5﹪CO 2 and cultured at 37°C. The medium was changed once every 2-3 days, and the culture was passaged once every 4-5 days. Well-grown OMC685-1 cells were digested with 0.25% trypsin to make a cell suspension, and 1×10 5 Inoculate in a 96-well culture plate at a density of 0.1 ml / well at 37°C, 5% CO 2 Culture in an incubator, and wait for the cells to adhere to the wall and cover the bottom of the well the next day;

[0023] (2) Infected cells and observing CPE with an inverted microscope

[0024] Rinse the 96-well culture plate three times repeatedly with PBS, blot the remaining PBS solution in the wells, and inoculate each well with throat...

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Abstract

The invention discloses a method for separating and culturing RSV (Respiratory Syncytial Virus) by using OMC (Ostiomeatal Complex)-685-1 cells, which comprises the following steps of: using the OMC-685-1 cells as a respiratory syncytial virus separating tool, observing a cytopathogenic effect by using an inverted microscope and detecting the virus by using an indirect immuno fluorescence method, and specifically comprises the following steps of: culturing the OMC-685-1 cells, infecting an OMC-685-1 cell line, observing a CPE (Cytopathic Effect) by using the inverted microscope and detecting the respiratory syncytial virus by using the indirect immuno fluorescence method. With the adoption of the separating and culturing method, whether a collected specimen contains RSV or not can be directly judged. The method has the advantages of simplicity, good repeatability, high sensitivity, short time consumption and low cost.

Description

technical field [0001] The invention relates to a method for isolating and culturing respiratory syncytial virus, in particular to a method for isolating and cultivating respiratory syncytial virus with OMC685-1 cells. Background technique [0002] Respiratory syncytial virus (RSV) belongs to the Pneumovirus genus of the Paramyxoviridae family, and has various shapes under the electron microscope, which can be ciliated or approximately spherical. [0003] RSV has poor tolerance to various factors in the environment, and high temperature, low pH, organic solvents, detergents, etc. can make it inactivated quickly. Therefore, it is required to inoculate sensitive cells as soon as possible during clinical isolation of viruses. RSV can grow in some passage cells, such as HEp-2, HeLa, HFF, A549 and rhesus monkey kidney cells. Under ideal conditions, RSV-specific confluent cells can be seen as early as two days after inoculation, but it usually takes 4-5 days for cytopathic effec...

Claims

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Application Information

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IPC IPC(8): C12N7/00G01N33/569
Inventor 敖弟书吴中明彭志元唐君肖福峰
Owner ZUNYI MEDICAL UNIVERSITY
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