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Hybridoma cell strain RSVN4C3 secreting anti-respiratory syncytial virus monoclonal antibody

A technology of hybridoma cell lines and monoclonal antibodies, which can be used in antiviral immunoglobulin, analytical materials, biomaterial analysis, etc., and can solve the problem of little research on N protein

Active Publication Date: 2021-08-13
GUIZHOU PROVINCIAL PEOPLES HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there are few studies on N protein in China at present.

Method used

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  • Hybridoma cell strain RSVN4C3 secreting anti-respiratory syncytial virus monoclonal antibody
  • Hybridoma cell strain RSVN4C3 secreting anti-respiratory syncytial virus monoclonal antibody
  • Hybridoma cell strain RSVN4C3 secreting anti-respiratory syncytial virus monoclonal antibody

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Experimental program
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Effect test

Embodiment 1

[0037] A hybridoma cell line RSVN4C3 secreting anti-respiratory syncytial virus monoclonal antibody, deposited in the China Center for Type Culture Collection (CCTCC), the address is Wuhan, China, the date of deposit is December 1, 2020, and its deposit number is CCTCC NO: C2020254.

[0038] The preparation method of this hybridoma cell line RSVN4C3 is:

[0039] Materials and Methods

[0040]The culture supernatant of RSV Guizhou 2017 isolated strain B133 Hep-2 cells was taken, the total RNA of RSV was extracted according to the instructions of Trizol reagent, and the first strand of cDNA was synthesized by reverse transcription. Using it as a template, specific primers were designed. The sequence is as follows:

[0041] P1: 5′-AAT GGATCC ATGGCTCTTTAGCAAAGTC-3' (the underlined part is the BamHI restriction site),

[0042] P2: 5′-ATT GTC GAC AAGCTTCATCATCATTATC-3' (the underlined part is the Sall restriction site);

[0043] Use P1 and P2 as primers to amplify the RSV N ge...

Embodiment 2

[0058] The monoclonal antibody against respiratory syncytial virus nucleocapsid protein secreted by hybridoma cell line RSVN4C3 was used to detect clinical respiratory syncytial virus infection by indirect immunofluorescence.

[0059] Take samples of nasopharyngeal secretions or nasopharyngeal washings from patients with respiratory tract infection, dilute them with normal saline, and centrifuge at 2000 rpm for 5 minutes to collect nasopharyngeal exfoliated cells. After suspending the exfoliated cells in 100 microliters of normal saline, drop the cells onto a glass slide , Acetone-fixed cells after drying. Add RSVN4C3 monoclonal antibody diluted 1:10000, dilute it with phosphate buffered solution (PBS) containing 3% bovine serum albumin (BSA), react at 37°C for 30 minutes, wash slides with PBS for 10 minutes, add 1:50 dilution FITC fluorescently labeled goat anti-mouse IgG antibody, reacted at 37°C for 30 minutes, washed the slide with PBS for 10 minutes, dried the slide, adde...

Embodiment 3

[0061] This embodiment provides a test kit for detecting or diagnosing respiratory syncytial virus infection, the test kit is an immune colloidal gold test strip, and the test kit includes the anti-respiratory syncytial virus nucleocoat secreted by the hybridoma cell line RSVN4C3 Monoclonal antibody to capsid protein.

[0062] The test strip includes a sample pad, a colloidal gold pad, a nitrocellulose membrane, an absorbent paper and a PVC bottom plate used as an assembly platform below in order of connection, and there is an overlap between the nitrocellulose membrane and the colloidal gold pad. There is overlap between the nitrocellulose membrane and the absorbent paper, and the colloidal gold pad is composed of a glass cellulose membrane adsorbed with a colloidal gold-labeled anti-RSVN4C3 monoclonal antibody, and the nitrocellulose membrane has goat anti-RSVN4C3 monoclonal antibody. The quality control line coated with mouse IgG polyclonal antibody, and the detection line ...

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Abstract

The invention discloses a hybridoma cell strain RSVN4C3 secreting an anti-respiratory syncytial virus monoclonal antibody. The hybridoma cell strain RSVN4C3 is preserved in China Center for Type Culture Collection (CCTCC), and the preservation number is CCTCC NO: C2020254. The anti-respiratory syncytial virus nucleocapsid protein monoclonal antibody secreted by the hybridoma cell strain RSVN4C3 is used for preparing a kit for detecting or diagnosing respiratory syncytial virus infection, and the kit can be an immune colloidal gold test strip or an ELISA kit. The hybridoma cell strain RSVN4C3 can stably secrete the anti-respiratory syncytial virus monoclonal antibody (MAb), the reactivity and specificity of the monoclonal antibody (MAb) are identified, and a foundation is laid for future diagnosis of RSV virus infection and research of an infection mechanism.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a hybridoma cell strain RSVN4C3 secreting monoclonal antibody against respiratory syncytial virus. Background technique [0002] Human respiratory syncytial virus (RSV) is the leading cause of acute lower respiratory infection (ALRI) worldwide, leading to hospitalization in children younger than 5 years and immunocompromised adults. RSV infection is associated with high morbidity and mortality from acute lower respiratory infections (ALRIs) in children in both developed and developing countries. In developed countries, it is the leading cause of infant hospitalization within one month of birth; in developing countries, RSV is the leading cause of infant death, and about 34 million children under the age of 5 develop severe RSV disease each year, of which 350 million children required hospitalization and between 66,000 and 199,000 died, 99 percent of which occurred in developing coun...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/20C07K16/10G01N33/577G01N33/569G01N33/558G01N33/58
CPCC07K16/1027G01N33/577G01N33/56983G01N33/558G01N33/587G01N2333/135G01N2469/10Y02A50/30
Inventor 余福勋查艳詹琳叶芝旭刘琳杨斌
Owner GUIZHOU PROVINCIAL PEOPLES HOSPITAL
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