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Acquired immune deficiency syndrome virus and treponema pallidum fusion protein as well as preparation method and application thereof

A technology of Treponema pallidum and HIV, which is applied in the field of immunological detection, can solve the problems of high total detection cost, time-consuming operation, inconvenient and other problems, and achieve the effect of low detection cost

Inactive Publication Date: 2011-09-07
中华人民共和国吉林出入境检验检疫局
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The existing commercialized colloidal gold test strips detect AIDS or syphilis antibodies separately. The operation is time-consuming and inconvenient, and most of them are imported reagents. and syphilis antibody test strips

Method used

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  • Acquired immune deficiency syndrome virus and treponema pallidum fusion protein as well as preparation method and application thereof
  • Acquired immune deficiency syndrome virus and treponema pallidum fusion protein as well as preparation method and application thereof
  • Acquired immune deficiency syndrome virus and treponema pallidum fusion protein as well as preparation method and application thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] Preparation of Recombinant Fusion Protein Antigen of HIV and TP Genetic Engineering

[0049] (1) Acquisition of major dominant antigen genes of HIV-1 / 2 and TP

[0050] The complete gene sequences of HIV-1 and type 2 viruses were searched in the Genebank database, and the gene sequences of the main dominant antigens gp41, gp120, gp36, and gp125 were determined, the recombinant gene templates were obtained by artificial synthesis, and a large number of genes were obtained by PCR. The HIV gene template PCR primers are as follows:

[0051] Sense primer:

[0052] GGATCC AGTTAAAATCGAACCGCTG

[0053] Anti-sense primer:

[0054] AGCTT TCA CAGGTCTTTCTTCAGAGATCAGTTTCTGTTC GCACGGCGCGTAGTTA.

[0055] TP gene template PCR primers are as follows:

[0056] Sense primer:

[0057] GC GGATCC ATGTGTTTCTTGCACCACTGT

[0058] Anti-sense primer:

[0059] GC GAATTC TCA CAGGTCTTTCTTCAGAGATCAGTTTCTGTTC CGCTTCTTTTTCACCACG

[0060] in GGATCC for BamHI Restriction sites, ...

Embodiment 2

[0070] Preparation of immune colloidal gold probes for recombinant antigens of HIV and TP genetic engineering:

[0071] 1) Preparation of colloidal gold Referring to the trisodium citrate reduction method and microwave technology, the optimal ratio of gold chloride and trisodium citrate for the preparation of 40 nm colloidal gold particles was determined by electron microscope observation. The operation steps are briefly described as follows: Boil 100 mL of 0.1 g / L gold chloride in a microwave oven, and quickly add 10 g of freshly prepared / L trisodium citrate, when the color of the solution turns dark blue, place it in a microwave oven and continue heating for 5 min, then make up for the lost water after cooling at room temperature, 25mM K 2 CO 3 The pH of the colloidal gold solution was adjusted, filtered through a 0.22 μm filter membrane and stored at 4 °C for later use. At the same time, the particle size and particle uniformity were observed under an electron microscope...

Embodiment 3

[0076] Preparation of GICA one-step dual rapid detection of AIDS and syphilis antibody test strips:

[0077] 1) Soak the glass cellulose membrane (300×20 mm) in 0.01 mol / L pH 7.4 phosphate buffer (20 g BSA, 25 g trehalose, 3 g PVP-40, 0.2 g NaN 3, 8 g NaCl, 0.2 g KCl, 2.9 g NaCl 2 HPO 4 12H 2 O,0.2 g KH 2 PO 4 , dilute to 1000 mL with ultrapure water for 30 min, and then dry at 37 °C to obtain a sample processing pad, which is vacuum-packed and stored at 4 °C for use.

[0078] 2) Colloidal gold-labeled two genetically engineered recombinant antigens, HIV and TP, were sprayed onto the glass cellulose membrane (300 mm×6 mm) at a volume of 50 μL per centimeter by the Bio-Dot film spotter to obtain gold-labeled conjugates Pad, after drying in a freeze dryer; paste it on the sample pad obtained in step 1);

[0079] 3) Immobilize HIV and TP two genetically engineered recombinant antigens and antibodies against HIV and TP two genetically engineered recombinant antigens on the p...

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Abstract

The invention provides acquired immune deficiency syndrome virus and treponema pallidum fusion protein which can be used for preparing GICA (Gold Immunochromatographic Assay) one-step double-combined acquired immune deficiency syndrome and syphilis antibody rapid detection test paper, simultaneously and jointly detecting acquired immune deficiency syndrome and syphilis viruses, simultaneously carrying out combined detection on two diseases and carrying out result reading by naked eyes without special instruments, thereby providing a convenient, rapid, accurate and cheap detecting method for the site rapid detection, such as blood center street compulsory blood donor screening, family self-help detection, disease screening for frontier port exit and entry people, and the like, site epidemiology investigation and other mechanisms needing rapid detection.

Description

technical field [0001] The invention discloses a fusion protein of AIDS virus and Treponema pallidum and a preparation method thereof, and further provides a GICA one-step dual-link rapid detection of AIDS and syphilis antibody test strips and a preparation method, which can rapidly detect AIDS and syphilis antibodies at the same time, and is suitable for on-site rapid detection Detection and epidemiological investigation, belonging to the field of immunological detection. Background technique [0002] AIDS and syphilis (hereinafter referred to as: HIV and TP) are infectious diseases that seriously endanger public health. The two diseases are transmitted through sexual transmission, mother-to-child transmission, and blood transmission. In recent years, the incidence of AIDS and syphilis has shown an upward trend. Early detection of cases can play a positive role in timely medical treatment and timely control. [0003] Colloidal gold immunochromatography (Gold Immuno-Chromat...

Claims

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Application Information

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IPC IPC(8): C07K19/00C12N15/70G01N33/531G01N33/571C12R1/19
Inventor 杨怀宁浦昀徐卉邵丽筠李娟柴景春邹红王振国谢兵赵大力王英超刘洋徐宏寇玲
Owner 中华人民共和国吉林出入境检验检疫局
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