Serum-free full-suspension culture type F81 cell line as well as construction method and application thereof

A suspended cell, full suspension technology, applied in the biological field, can solve the problem of non-existence, and achieve the effect of reducing the probability of pollution, considerable economic benefits, and guaranteed safety.

Active Publication Date: 2022-04-12
广东省华晟生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, there is no serum-free...

Method used

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  • Serum-free full-suspension culture type F81 cell line as well as construction method and application thereof
  • Serum-free full-suspension culture type F81 cell line as well as construction method and application thereof
  • Serum-free full-suspension culture type F81 cell line as well as construction method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0054] Example 1 Construction of Serum-Free Full Suspension Culture Type F81 Cell Line

[0055] 1. Resuscitate an adherent cultured F81 cell, inoculate it into the MEM medium containing 10% fetal bovine serum (take MEM (Gibco company product) as the base medium, add 10% fetal bovine serum (Gibco company product) to obtain) cultured at 37°C, 5% CO 2 , pH7.2; after the cells adhered to the wall and the confluence of the monolayer in the culture flask reached 95%, the culture medium was completely discarded, digested and dispersed with 0.25% EDTA-trypsin, subcultured for 3 generations according to the conventional method, the cell edge was clear, Flat shape, normal growth can be used for domestication;

[0056] 2. Culture the adherent cultured F81 cells obtained in step 1 with MEM medium containing 6%, 4%, and 2% fetal bovine serum for 3 generations, and the passage ratio is 1:3, and the culture conditions are as follows: 37°C, 5% CO 2 , pH7.2, to obtain low serum adherent F81...

Embodiment 2

[0061] Example 2 Inspection of Serum-Free Full Suspension Culture Type F81 Cell Line

[0062] 1. Sterility test

[0063] Get the supernatant of the serum-free F81 cell culture process obtained in step 5 of Example 1 and inoculate them into thioglycolate medium, caseptone agar medium (slant) and glucose-peptone medium and culture them respectively (respectively at 37° C. And room temperature (22 ~ 26 under cultivation, each group of 5 tubes of cell sample solution, observed for 7 days), the results were negative, indicating sterile contamination.

[0064] 2. Growth characteristics of F81 cells at different seeding densities

[0065] 5×10 5 cells / mL, 10×10 5 cells / mL, 15×10 5 cells / mL Serum-free full suspension culture F81 cells were inoculated and cultured at 37°C, 5% CO 2 And 120r / min shaker culture, every 24h sampling observation and counting, the results are as follows: at 5~10×10 5 The cell viability in the range of cells / mL inoculation is maintained above 90%, and th...

Embodiment 3

[0066] Embodiment 3 The characteristics of serum-free full suspension culture type F81 cell line

[0067] The adherent cultured F81 cells before domestication, the low-serum adherent F81 cells obtained in step 2 of Example 1, and the 13th generation serum-free full suspension cultured F81 cells were inoculated into the corresponding medium (the adherent culture before domestication). Cultured F81 cells are MEM medium with 5% fetal bovine serum concentration, low serum adherent F81 cells are MEM medium with 2% fetal bovine serum concentration, serum-free full suspension culture type F81 cells are MS01 medium), 37 Cultivate for 48 hours under 5% carbon dioxide concentration, the results are as follows figure 1 , 2 , as shown in 3: figure 1 It shows that the adherent cultured F81 cells before acclimatization adhere to the surface of the culture medium in the state of adherent culture with serum, showing the shape of mosaic paving stones; figure 2 It shows that the low-serum a...

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Abstract

The invention belongs to the technical field of biology, and discloses a serum-free full-suspension culture type F81 cell line as well as a construction method and application thereof. The cat kidney suspension cell line is named as a cat kidney cell F81 suspension cell, is preserved in China Center for Type Culture Collection in Wuhan University, Wuhan, China on November 4, 2021, has a preservation number of CCTCC NO: C2021289, has been passed to 65 generations at present, and is mainly in a transparent round shape with uniform size; a virus sensitivity test shows that the serum-free full-suspension culture type F81 cell line is sensitive to feline parvovirus; the cell line can be used for culturing, separating and detecting viruses, preparing viral vaccines and screening drugs for preventing or treating diseases caused by virus infection.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a serum-free full suspension culture type F81 cell line and a construction method and application thereof. Background technique [0002] Feline panleukopenia (FPL), also known as feline distemper, is a highly contagious disease caused by Carnivore protoparvovirus 1. The typical symptoms are panleukopenia, severe enteritis and Immunosuppression, with high morbidity and mortality. So far, no ideal treatment drug has been found for feline distemper, and vaccination is still the most effective means to prevent the occurrence and prevalence of the disease today. [0003] With the continuous development of the pet industry, the number of pets is increasing year by year, mainly pet cats and pet dogs. While pets bring happiness to people, pet health problems also threaten public safety. At present, the viruses that harm pet cats mainly include feline parvovirus, canine parvovir...

Claims

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Application Information

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IPC IPC(8): C12N5/071C12N7/00C12Q1/04C12Q1/70C12Q1/02A61K39/23A61P31/20C12R1/93
CPCY02A50/30
Inventor 朱辉张久鑫张梓鸿邵宝顺盛圆贤
Owner 广东省华晟生物技术有限公司
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