Antigen pre-processing method for preparing antibody

A pretreatment and antigen technology, applied in the field of immunology, can solve the problems of poor specificity, inaccurate antibody positioning, lack of pertinence and purpose of antigen, and achieve the effect of low production cost and easy operation.

Inactive Publication Date: 2015-07-01
FUZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The antigen obtained by only simple protein purification steps lacks pertinence and purpose, and the antibody prepared by using it as an immunogen often has inaccurate antibody positioning and poor specificity in the application of immunohistochemical techniques
In view of this, most of the high-quality antibodies that are well used in WB and ELISA techniques cannot be applied to immunohistochemical techniques

Method used

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  • Antigen pre-processing method for preparing antibody
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  • Antigen pre-processing method for preparing antibody

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Add 0.01 mol / L EDTA buffer solution with pH values ​​of 7.0, 8.0, 9.0, and 10.0 to the antigen protein solution 1 (membrane protein), respectively, the final concentration of the antigen protein solution 1 is 1 mg / mL, and treat at 100°C for 20 minutes.

[0021] The mice were immunized with the antigen retrieval solution treated and untreated antigen protein respectively. After three times of immunization, the blood antibody titer was detected by ELISA, and the detection titer reached 10 6 , Dilute the antibody 1000 times with antibody diluent and perform immunohistochemical experiments ("Immunohistochemical Test Manual"). In this experiment, mice immunized with untreated antigen protein were used as negative control, and blank mice without immunization were used as blank control. The results of immunohistochemical staining were as figure 1 shown.

[0022] Depend on figure 1 It can be seen that the blank control in Figure A is the negative result of immunohistochemist...

Embodiment 2

[0024] Add 0.01 mol / L citrate buffer with pH values ​​of 4.0, 5.0, 6.0, and 7.0 to the antigenic protein solution 2 (membrane protein) respectively, the final concentration of the antigenic protein is 1mg / mL, and treat at 120°C for 90s.

[0025] The mice were immunized with the antigen retrieval solution treated and untreated antigen protein respectively. After three times of immunization, the blood antibody titer was detected by ELISA, and the detection titer reached 10 6 , Dilute the antibody 1000 times with antibody diluent and perform immunohistochemical experiments. In this experiment, mice immunized with untreated antigen protein were used as negative control, and blank mice without immunization were used as blank control. The results of immunohistochemical staining were as figure 2 shown.

[0026] Depend on figure 2 It can be seen that the blank control in Figure A is the negative result of immunohistochemistry, and the immunohistochemical staining results of the n...

Embodiment 3

[0028] Add 0.01 mol / L Tris buffer with pH values ​​of 6.0, 7.0, 8.0, and 9.0 to the antigenic protein solution 3 (membrane protein), and the final concentration of the antigenic protein solution 3 is 1 mg / mL, and treat at 100°C for 20 minutes.

[0029] The mice were immunized with the antigen retrieval solution treated and untreated antigen protein respectively. After three times of immunization, the blood antibody titer was detected by ELISA, and the detection titer reached 10 6 , Dilute the antibody 1000 times with antibody diluent and perform immunohistochemical experiments. In this experiment, mice immunized with untreated antigen protein were used as negative control, and blank mice without immunization were used as blank control. The results of immunohistochemical staining were as image 3 shown.

[0030] Depend on image 3 It can be seen that the blank control in Figure A is the negative result of immunohistochemistry, and the immunohistochemical staining results of ...

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Abstract

The invention discloses an antigen pre-processing method for preparing an antibody, and relating to the technical field of immunology. Purified antigen protein is processed by adopting an antigen retrieval solution in the pre-processing method. Aiming at the characteristics of an immunohistochemistry technology and disadvantages in the prior art, the invention aims to provide the antigen pre-processing method; and the antigen pre-processing method has the advantages of being simple and convenient to operate, low in production cost and the like. The key step of immunohistochemistry in the invention is antigen retrieval; the antigen is processed by selecting a corresponding antigen retrieval solution; the antigen pre-processing method, which can be applied to immunohistochemistry, is designed in a targeted manner, i.e., the immunohistochemistry retrieval manner of the antibody is selected according to the processing manner of the antigen; the antigen pre-processing method is simple and convenient to operate and low in production cost; and a novel effective way is provided for preparing the antibody applied to immunohistochemistry.

Description

technical field [0001] The invention discloses an antigen pretreatment method for preparing antibodies and relates to the technical field of immunology. Background technique [0002] Immunohistochemical technology is a methodology developed by Sterb Berger in the early 1970s on the basis of enzyme labeling method and immunological antigen-antibody reaction as the theoretical basis. Protein) is a technology for localization, qualitative or quantitative detection. Immunohistochemical technology plays an extremely important role in the field of biology, especially in basic medical and clinical research. The introduction of its technology has changed the depth of research on tumor diagnosis and classification, making it from the cellular level to the biochemical level. , molecular level, greatly improving the accuracy and sensitivity of tumor diagnosis, and laying a good foundation for tumor treatment. Immunohistochemical technology is currently mainly used in clinical testin...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/00C07K1/00
Inventor 孟春郑美云王航陈小媛
Owner FUZHOU UNIV
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