Preparation method and application method of chlamydia pneumoniae recombinant antigen

A technology of Chlamydia pneumoniae and recombinant antigen, which is applied in the field of medical drugs, can solve the problems of infection, slow detection speed and low accuracy of preparation personnel, and achieves the effects of low cost, simple operation and good stability.

Inactive Publication Date: 2020-05-01
南京拂晓生物科技有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] The purpose of the present invention is to provide a preparation method of Chlamydia pneumoniae recombinant antigen and its application method to solve practical problems such as slow detection speed, low accuracy, cumbersome process, high cost and risk of infection for preparation personnel at the present stage

Method used

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  • Preparation method and application method of chlamydia pneumoniae recombinant antigen
  • Preparation method and application method of chlamydia pneumoniae recombinant antigen
  • Preparation method and application method of chlamydia pneumoniae recombinant antigen

Examples

Experimental program
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Effect test

Embodiment 1

[0035] see figure 1 As shown, a preparation method of Chlamydia pneumoniae recombinant antigen, the specific steps are as follows: (1) Selection of Chlamydia pneumoniae recombinant antigen sequence: comprehensively compare the antigen targets reported in the existing literature of CP, and combine bioinformatics analysis to finally determine the incA , the most immunogenic amino acid segments of incB and opmA are: amino acid 220-amino acid 490 of incA, amino acid 98-amino acid 175 of incB gene, amino acid 25-amino acid 175 of ompA gene Amino acids at position 192 are connected in series through the connecting peptide GSGSGS. The series sequence from N-terminal to C-terminal is incA fragment-GSGSGS-incB fragment-GSGSGS-ompA fragment. The final nucleic acid sequence is obtained by chemical synthesis.

[0036] See SEQ ID NO.1 for the nucleotide sequence of the CP recombinant antigen, and SEQ ID NO.2 for the amino acid sequence. After chemically synthesizing the target gene fragmen...

Embodiment 2

[0047] see figure 2 , 3 Shown, a kind of application method of Chlamydia pneumoniae recombinant antigen, its specific steps are as follows:

[0048] (1) Dilute the CP protein to 2 μg / mL with coating buffer (0.1 mol / L carbonate buffer at pH 9.6), add it to the well of the microtiter plate, 100 μL / well, incubate at 37°C for 2 hours; shake off After coating, pat dry, wash with PBST 3 times, 3min / time; add 200μL of blocking solution (containing 2% bovine serum albumin phosphate buffer at final concentration) to each well, incubate at 37°C for 2h; shake off the bag After soaked, pat dry, and wash 3 times with PBST, 3min each time; at this time, it can be stored in vacuum packaging in aluminum foil bags;

[0049] (2) Dilute the sample to be tested 1:100 with the sample diluent and mix well;

[0050] (3) Add 100 μL of diluted sample to each well, and add 100 μL of CP-IgM calibrator solution to the reserved calibrator wells. After sealing the wells of the microplate plate, incubat...

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Abstract

The invention discloses a preparation method and an application method of a chlamydia pneumoniae (CP) recombinant antigen, wherein the preparation method of the CP recombinant antigen comprises the following specific steps: selection of CP recombinant antigen sequences; prokaryotic expression of the CP recombinant antigen; purification and renaturation of CP recombinant antigen; and verification of CP recombinant antigen. A serological detection method is developed on the premise of preparing an effective CP antigen, a preparation process of a CP natural antigen is complicated and high in cost, and there is a risk of infection for preparation personnel. Using the recombinant antigen with strong immunogenicity and good specificity as a substitute of the natural antigen not only reduces production cost, but also lays a foundation for developing a detection kit for detecting CP antibodies, IGG and IgM. The method is simple in operation and low in cost, the CP antigen is strong in specificity and good in stability, capable of achieving mass production and suitable for popularization and application.

Description

technical field [0001] The invention relates to a medical drug, in particular to a preparation method and an application method of a Chlamydia pneumoniae recombinant antigen. Background technique [0002] Chlamydia pneumonia (CP) is one of the main pathogens of community-acquired pneumonia, which is relatively common in the world. Young children and the elderly are susceptible groups. The disease can occur in all seasons, and it is mainly transmitted through droplets. Clinically, the main symptoms are respiratory symptoms, sore throat, hoarseness, runny nose, etc., and fever and cough may also be manifested. CP infection is usually a mild, self-limiting disease that is often overlooked or underdiagnosed because of its atypical symptoms. Studies have shown that CP infection is related to some chronic diseases, such as asthma, otitis media, erythema nodosum, atherosclerosis and endocarditis, coronary heart disease, etc. CP infection can also cause damage to the cerebrovascula...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/295C12N15/70G01N33/68G01N33/569
CPCC07K14/295C12N15/70G01N33/68G01N33/56927G01N2333/295
Inventor 李伟郭晓
Owner 南京拂晓生物科技有限公司
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