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Test paper for detecting borrelia burgdorferi antibodies, test paper preparation method and kit

A technology of Borrelia burgdorferi and a kit, which is applied in the direction of resisting vector-borne diseases, measuring devices, instruments, etc., to achieve the effects of strong specificity, low reagent cost, and accurate and reliable test results

Active Publication Date: 2015-02-04
CHINESE ACAD OF INSPECTION & QUARANTINE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there is no report of a recombinant protein that can be effectively used as an antigen of Borrelia burgdorferi

Method used

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  • Test paper for detecting borrelia burgdorferi antibodies, test paper preparation method and kit
  • Test paper for detecting borrelia burgdorferi antibodies, test paper preparation method and kit
  • Test paper for detecting borrelia burgdorferi antibodies, test paper preparation method and kit

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Experimental program
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Embodiment 1

[0049] The preparation of embodiment 1 Borrelia burgdorferi antigen

[0050] The specific steps for preparing the Borrelia burgdorferi antigen are as follows:

[0051] a. Construction of recombinant plasmids

[0052] To the selection of Borrelia burgdorferi antigen, the inventor of the present invention carries out multiple tests, finally selects Borrelia burgdorferi envelope OSPA protein, its amino acid sequence reference (GenBank sequence number: CP001318.1), according to Escherichia coli Escherichia coli O127: H6 preferred codons were used to modify the gene sequence, and the secondary structure of the recombinant protein was screened through bioinformatics (see the reference materials for substantive review of the screening scheme, which will not be repeated here), and after multiple experiments, the final gene sequence was determined As shown in SEQ ID NO.1.

[0053] According to the sequence shown in SEQ ID NO.1, entrust Shanghai Yingjun Company to synthesize it, conne...

Embodiment 2

[0059] Embodiment 2 detects the preparation of the test paper of Borrelia burgdorferi antibody

[0060] A. SPA labeling magnetic nanoparticles, and preparing magnetic nanoparticle carrier pads:

[0061] Take 100 μl ferroferric oxide magnetic nanoparticle solution (the particle size of ferric oxide tetroxide is preferably 12nm), add 700 μl water, 200 μl 25% glutaraldehyde solution, shake for 3 h; wash 1-2 times with 1 ml water, wash 2 times in PBS The second time is to carry out carboxylation treatment on ferric ferric oxide magnetic nanoparticles; add 500 μl of 2 mg / ml SPA, shake for 3 hours, and the pH value is 7.6; add 500 μl of 0.5% BSA, shake for 30 minutes, and block; add 0.01% spit Warm-20 PBS with a concentration of 0.01M, wash 3-4 times; add 1mL resuspension solution, wherein the resuspension solution contains 0.1% Tris base, 1% BSA and 5% sucrose by weight percentage, mix well, Spray on glass fiber membrane and dry at 37°C for 4 hours;

[0062] B, coated nitrocellul...

Embodiment 3

[0067] The detection of the test paper of the detection Borrelia burgdorferi antibody prepared by the present invention and the sensitivity of colloidal gold immunochromatography test paper

[0068]Wherein, the method for the test paper of the detection Borrelia burgdorferi antibody prepared by the present invention sees embodiment 2;

[0069] The preparation method of colloidal gold immunochromatography test paper is as follows:

[0070] Colloidal gold immunochromatography test paper was prepared by conventional methods, wherein SPA-labeled colloidal gold particles were used, the detection zone on the nitrocellulose membrane was coated with Borrelia burgdorferi recombinant antigen, and the quality control zone was coated with goat anti-rabbit IgG.

[0071] The test strips prepared in Example 2 were used to detect respectively the Borrelia burgdorferi antibody serum, diluted to 1:10, 1:100, 1:1000, 1:10000, 1:100000, 1:1000000 with fetal bovine serum, and simultaneously Fetal...

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Abstract

The invention discloses a piece of test paper for detecting borrelia burgdorferi antibodies, a test paper preparation method and a kit. The invention discloses the test paper for detecting the borrelia burgdorferi antibodies. The test paper consists of a sample pad, a Fe3O4 magnetic nano particle carrier pad, a nitrocellulose film and a water absorption pad which are overlapped in sequence and are attached to a bottom lining clamp, wherein the Fe3O4 magnetic nano particle carrier pad is a glass fiber film on which staphylococcus aureus A protein (SPA) labeled magnetic nano particles are fixed; the nitrocellulose film is provided with a detection band and a quality control band; the detection band is wrapped by borrelia burgdorferi antigens; the quality control band is wrapped by antibodies combined with SPA. The detection kit consists of the detection test paper and peroxidase substrate developing liquid and is used for detecting the borrelia burgdorferi antibodies. The kit can effectively detect the borrelia burgdorferi antibodies; a detection result is stable and reliable; the specificity is high, and the sensitivity is high; compared with a piece of general colloid detection test paper, the test paper has the advantages that the test paper sensitivity is improved by 100 times, and the operation is simple; detectors are not needed; the detection result can be obtained by eyes.

Description

technical field [0001] The invention relates to the field of biological detection reagents, in particular to a test paper for detecting Borrelia burgdorferi antibodies, a method for preparing the test paper and a kit. Background technique [0002] At present, the detection technology for Borrelia garinii antibody is mainly isolation culture and serological experiment. Because the isolation culture takes a long time, immunological methods such as ELISA and colloidal gold are generally used. Among them, the ELISA detection method needs repeated cleaning, The decision was made, and the absorbance detection was required by the scenery photometry instrument, and it took a long time; the colloidal gold detection method was suitable for on-site detection, but the detection sensitivity was limited, mainly because the colloidal gold was connected with the antibody or protein to be labeled. Stable, it is easy to detach the antibody or labeled protein due to the change of charge, resul...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/558G01N33/544
CPCG01N33/544G01N33/558Y02A50/30
Inventor 杨宇徐宝梁曾婷婷赵婷婷王静
Owner CHINESE ACAD OF INSPECTION & QUARANTINE
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