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Meningococcal capsular polysaccharide monoclonal antibody and its application

A monoclonal antibody, meningococcal technology, applied in the direction of antibacterial immunoglobulins, instruments, biochemical equipment and methods, etc., can solve the problem of ELISA kits for the rapid and effective detection of meningococcal capsular polysaccharides, etc. question

Active Publication Date: 2018-04-03
ABMAX BIOTECHNOLOGY CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] At present, there are no reports of ELISA kits for rapid and effective detection of meningococcal capsular polysaccharides at home and abroad.

Method used

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  • Meningococcal capsular polysaccharide monoclonal antibody and its application
  • Meningococcal capsular polysaccharide monoclonal antibody and its application
  • Meningococcal capsular polysaccharide monoclonal antibody and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] Embodiment 1, establishment of hybridoma cell line

[0046] 1. Experimental materials

[0047] 1. Immunogen: The capsular polysaccharides of meningococci of groups A, C, Y, and W135 were used as immunogens (purchased from Tianjin CanSino Biotechnology Co., Ltd.).

[0048] 2. Culture medium: DMEM medium was purchased from Hyclone Company; HAT, HT selection medium and pristane were purchased from Sigma Company.

[0049] 3. Experimental animals: Balb / c mice, 8-12 weeks old, female, SPF grade animal culture.

[0050] 4. Other materials: Freund's complete adjuvant and Freund's incomplete adjuvant were purchased from Sigma Company; PEG4000 was purchased from Fluka Company; HRP-goat anti-mouse IgG antibody was purchased from JacksonImmune Company; other reagents were domestic analytically pure products.

[0051] 2. Establishment of hybridoma cell lines

[0052] 1. Animal immunity

[0053] 1) Basic immunization: The immunogen was mixed with Freund's complete adjuvant in equ...

Embodiment 2

[0065] Example 2 Preparation of Monoclonal Antibody Against ACYW135 Group Meningococcal Capsular Polysaccharide and Carrier Protein

[0066] Primary antibody preparation

[0067] The following antibody preparations were performed against hybridoma cells that can secrete monoclonal antibodies against A, C, Y, and W135.

[0068] Adult BALB / c mice were selected, and pristane was inoculated intraperitoneally, 0.5ml per mouse. After 7-10 days, the 16th generation hybridoma cells (hybridoma cells capable of secreting monoclonal antibodies against A, C, Y, and W135 were inoculated respectively) were inoculated intraperitoneally, 1×10 per mouse 6 -2×10 6 indivual. After an interval of 5 days, when the abdomen is obviously enlarged and the skin feels tense when touched with hands, the ascites can be collected with a No. 9 needle.

[0069] Centrifuge the ascitic fluid (13000r / min for 30 minutes), remove cell components and other precipitates, and collect the supernatant. Purify wit...

Embodiment 3

[0086] Example 3 Preparation of ACYW135 Group Meningococcal Capsular Polysaccharide Detection Reagent Using Purified Antibody

[0087] 1. Monoclonal antibodies Clone6B7 and Clone 6B12 for meningococcal A polysaccharide recognition by meningococcal A polysaccharide ELISA double antibody sandwich method.

[0088] Using clone 6B7 and Clone6B12 antibodies for pairing experiments, it was determined that clone 6B12 was used as the coating antibody, and HRP-labeled Clone 6B7 was used as the detection antibody. The ELISA detection method was determined, and the detection sensitivity of the kit could reach 0.008ng / mL ( image 3 ). Antibodies were labeled with the modified sodium periodate method.

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Abstract

The invention provides monoclonal antibodies to meningococcal capsular polysaccharides and applications thereof, respectively providing mouse monoclonal antibodies specific to polysaccharides of meningococcal groups A, C, Y and W135, by immunizing mice and preparing hybridomas cells obtained. The various types of meningococcal polysaccharide-specific monoclonal antibodies provided by the invention have no obvious cross-reaction with other types of capsular polysaccharides, and have the advantages of high specificity and high sensitivity for detecting meningococci of groups A, C, Y and W135. The method can accurately detect the contents of various types of meningococci or capsular polysaccharides in samples, and has broad application prospects in clinical testing and vaccine production.

Description

technical field [0001] The present invention relates to a hybridoma cell line and the monoclonal antibody produced by it and its application, in particular to the monoclonal antibody against the four capsular polysaccharides of Neisseria meningitidis A, C, W135, and Y serotypes and the hybridization of the antibody secreted Tumor cell lines and the application of the kit containing the antibody. Background technique [0002] Epidemic cerebrospinal meningitis (meningococcal meningitis) has been discovered for more than 100 years, and it is still prevalent in many countries. It is also a common acute respiratory infectious disease in winter and spring in my country. The disease is meningitis caused by the bacterium Neisseria meningitides. According to statistics, from February to April every year, the incidence of "Emergency Meningitis" accounts for about 60% of the whole year. Patients are mainly children under the age of 15, especially infants and young children aged 6 mon...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K16/12C12N5/20G01N33/577G01N33/535G01N33/569C12R1/91
Inventor 孙乐李茂华王慕旸张翠娟张小刚陈兴张萍萍
Owner ABMAX BIOTECHNOLOGY CO LTD