Preparation method of monoclonal antibody for replacing anti-mouse rabbit secondary antibody

A monoclonal antibody and anti-pika secondary antibody technology, which is applied in chemical instruments and methods, instruments, analytical materials, etc., can solve the problems of high cost and poor stability of the second antibody

Active Publication Date: 2020-04-10
SHANDONG LVDU BIO SICIENCE & TECH
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] For this reason, the embodiment of the present invention provides a method for preparing a monoclonal antibody used to replace the anti-pika secondary antibody, so as to solve the problems of poor stability and high cost of the second antibody in the prior art

Method used

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  • Preparation method of monoclonal antibody for replacing anti-mouse rabbit secondary antibody
  • Preparation method of monoclonal antibody for replacing anti-mouse rabbit secondary antibody
  • Preparation method of monoclonal antibody for replacing anti-mouse rabbit secondary antibody

Examples

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Embodiment 1

[0019] Example 1. Preparation of monoclonal antibody used to replace anti-pika secondary antibody

[0020] 1. Preparation of whole antigen immunogen from immune serum

[0021] (1) Wash 1ml of SPA agarose gel prepacked column (Shandong Lvdu Biotechnology Co., Ltd., LD-CH-02) with 5-10 column bed volumes of distilled water;

[0022] (2) Equilibrate 5 to 10 column bed volumes with binding buffer binding buffer (0.02M phosphate buffer, 0.01M NaCl, pH 7.4);

[0023] (3) Dilute 5ml of rabbit serum to 50ml with binding buffer, filter and load with 0.45μm filter membrane;

[0024] (4) Wash 5 to 10 column bed volumes with binding buffer;

[0025] (5) Elute the antibody with elution buffer (0.1M citric acid buffer, pH 4.0), collect the elution peak, and adjust its pH to neutral with neutralization buffer (1M Tris-HCl, pH 9.0), determine Protein concentration, used as an immunogen for backup;

[0026] (6) After each use, regenerate and wash with 3-5 column bed volumes of 0.1M NaOH so...

Embodiment 3

[0043] Example 3, the application of the monoclonal antibody used to replace the anti-pika secondary antibody in the immunochromatographic detection kit

[0044] 1. Application of the monoclonal antibody of the embodiment of the present invention in the qualitative detection card of chloramphenicol

[0045] 1.1 Labeling of chloramphenicol-binding protein

[0046] With 0.1 mol / L of K 2 CO 3Adjust the pH of the colloidal gold to 9.0, add 6 μg of chloramphenicol-binding protein to each 1 mL of colloidal gold solution, shake at 120 rpm for 30 minutes at room temperature, add 20 μL of 10% bovine serum albumin, shake at 120 rpm for 30 minutes at room temperature, centrifuge at 12,000 rpm for 20 minutes, discard the supernatant, The precipitate was dissolved with 0.01MPBS to obtain the labeled chloramphenicol-binding protein.

[0047] 2. Determination of quality control C line and detection line T line

[0048] Select the Sartorius Cn140 membrane, respectively draw the whole anti...

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Abstract

The embodiment of the invention discloses a preparation method of a monoclonal antibody for replacing an anti-mouse rabbit secondary antibody. The method comprises the steps of using purified rabbit serum as a holoantigen, and carrying out immunoreaction and cell fusion screening to obtain hybridoma cells. The embodiment of the invention provides the preparation method of a mouse non-specific monoclonal antibody, which uses the rabbit serum holoantigen as an immunogen, and obtains the anti-mouse rabbit secondary antibody used for replacing the prior art through immunogen purification, immune procedure, cell fusion, hybridoma screening and cloning, and antibody preparation and purification. The monoclonal antibody prepared by the embodiment of the invention is simple in preparation method,can be produced in batches, is good in stability, can be applied to immunochromatographic detection, is high in titer and strong in reaction, and is suitable for immunochromatographic reactions in various modes.

Description

technical field [0001] The embodiment of the present invention relates to the technical field of antibody preparation, and specifically relates to a preparation method of a monoclonal antibody used to replace the anti-pika secondary antibody. Background technique [0002] Antibodies from primary sources such as mice or rabbits are called primary antibodies, and secondary antibodies are antibodies that can bind to the primary antibody, that is, antibodies, whose main function is to detect the presence of antibodies and amplify the signal of the primary antibody. The secondary antibody is to use the antigenic property of the protein, which is a macromolecule, to immunize the xenogeneic animal, and the immunoglobulin produced by the immune system of the xenogeneic animal is directed against this antibody. The secondary antibody has played a huge role in experiments such as indirect ELISA and immunochromatography. Immuno-chromatography assay (ICA) is a new type of immunoassay m...

Claims

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Application Information

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IPC IPC(8): C07K16/42G01N33/577G01N33/558
CPCC07K16/42G01N33/558G01N33/577
Inventor 武玉香沈志强于金枝崔平
Owner SHANDONG LVDU BIO SICIENCE & TECH
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