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Detection card for quickly quantitative detection of echinococcosis antibody in serum

A quantitative detection and antibody detection technology, which is applied in the field of detection cards, can solve the problems of narrow detection linear range, low sensitivity, and only qualitative detection, and achieve the effect of improving sensitivity, high sensitivity, and small intra-assay difference

Inactive Publication Date: 2018-04-06
杭州微瑞科技有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

ELISA kits for the detection of echinococcosis antibodies are also used on the market. ELISA kits require a microplate reader, incubation reaction conditions, plate washing conditions, operating environment and professional technicians. In addition, detection samples also require complex pretreatments, such as Collect blood, separate serum, etc.; while rapid colloidal gold detection is convenient and does not require professional technicians and working environment, but its detection sensitivity is low, and it can only be qualitative, and the detection linear range is narrow

Method used

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  • Detection card for quickly quantitative detection of echinococcosis antibody in serum
  • Detection card for quickly quantitative detection of echinococcosis antibody in serum
  • Detection card for quickly quantitative detection of echinococcosis antibody in serum

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Embodiment 1, echinococcosis Eg95 protein prokaryotic expression

[0037] Step 1. Construction of echinococcosis antigen gene recombinant expression plasmid pET-30a-(Eg95)

[0038] According to the echinococcosis Eg95 gene on the NCBI website, the target sequence of the Eg95 gene was optimized to the preferred codon sequence of Escherichia coli, and the Eg95 expression target gene sequence of the echinococcosis was synthesized by chemical synthesis method. The pET30a vector was digested with BamH I and Xho I. The digestion system was 40 μL: 4 μl of 10× digestion buffer, 1 μL of Ned I and Xho I, 24 μL of vector, 8 μL of deionized water, and the digested product was treated with 1% Gel recovery kit recovery after agarose electrophoresis. Then the digested vectors were ligated with the amplified Eg95 coding gene sequence to construct a 10 μL ligation system. Among them, 5.5 μL of Eg95 encoding gene fragment, 1.5 μL of pET30a vector, 1 μL of T4 DNA ligase, and 2 μL of T4 ...

Embodiment 2

[0044] Example 2, Eg95 protein epitope antigen expression of echinococcosis

[0045] A method for preparing the above echinococcosis epitope antigen, using bioinformatics technology to compare and analyze Eg95 antigen genes of different lineages of echinococcosis, design and synthesize the main epitope antigen gene of echinococcosis Eg95, in order to ensure the epitope antigen gene The expression vector was correctly inserted, and the specific enzyme cutting site EcoRI / xhoI was introduced at both ends of the gene. The gene was entrusted to Bao Bioengineering (Dalian) Co., Ltd. to synthesize it and named it AE. The synthetic gene AE and the recombinant expression plasmid pGEM-6P-1 were digested with EcoRI / xhoI enzymes, and the gene fragment and the DNA fragment of pGEM-6P-1 were recovered with a DNA fragment recovery kit (Takara, Dalian). Then use T4 ligase to ligate the two fragments to construct the recombinant plasmid pGEM-AE. The ligation product was transformed into BL21(...

Embodiment 3

[0047] Example 3: Preparation of Echinococcosis Antibody Detection Card

[0048] Preparation of echinococcosis antibody detection standard curve: prepare 6 copies of calibration solution containing echinococcosis antibody (including echinococcosis antibody standard), the concentrations are 0, 1 / 1024, 1 / 256, 1 / 64, 1 / 16 , 1 / 4 (double dilution of echinococcosis antibody standard). Add the above-mentioned calibration solutions of different concentrations into the sample holes of the assembled test card, and after 15 minutes of chromatography, the test is carried out by a tomographic scanner, and the test results obtained 6 times are processed by the client, and the client calculates The fluorescence signal intensity values ​​of the detection line and quality control line corresponding to the standard product, and perform linear regression based on this data to make a standard curve of echinococcosis antibody. The standard curve calculated by the client forms a file and generates a...

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Abstract

The invention discloses a detection card for quickly quantitative detection of echinococcosis antibody in serum. The detection card includes a detection card case and a test paper strip assembled therein. The test paper strip includes a plastic base board provided with pressure-sensitive adhesive, wherein a sample pad, a marker pad, a nitrocellulose membrane and water absorption paper are successively adhered to the base board. The marker pad is composed of a carrier base layer and a marker. The marker is a film formed by spray-coating the carrier base layer with lanthanum-series fluorescencedetection microspheres and lanthanum-series fluorescence quality control microspheres. The nitrocellulose membrane is coated with an echinococcosis recombinant antigen to form a detection line and rabbit-anti-chicken IgY antibody to form a quality control line. The marker includes the fluorescence detection microspheres labeled by the echinococcosis recombinant antigen and the fluorescence qualitycontrol microspheres labeled by the chicken IgY antibody. The detection card can achieve quick quantitative detection of the echinococcosis antibody in situ and has great practical and application values.

Description

technical field [0001] The invention relates to a detection card and its use, in particular to a detection card for rapid and quantitative detection of echinococcosis antibody in serum. Background technique [0002] Hydatidosis, also known as Echinococcus, is a zoonotic parasitic disease that seriously endangers the health of humans and animals. The Ministry of Agriculture lists it as a second-class animal disease, and the Ministry of Health lists it as a C class of infectious diseases. In my country, 25 provinces, municipalities, and autonomous regions have confirmed cases of infection, among which Gansu, Xinjiang, Ningxia, Inner Mongolia, Qinghai, Sichuan, Shaanxi, Tibet and other northwestern provinces are particularly serious. With the development of modern society, the floating population has increased, and the number of pets and commercial animals has increased. The circulation of animals and people has caused echinococcosis to gradually spread to inland and coastal p...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/559G01N33/561C12N15/70C12N15/12
CPCC07K14/43554C12N15/70G01N33/559G01N33/561
Inventor 吴俊清章健吴冠英王泽洲
Owner 杭州微瑞科技有限公司
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