Rapid quantitative detection card for canine distemper virus antibody and using method

A canine distemper virus, quantitative detection technology, applied in the field of detection cards, can solve the problems of narrow detection range, only qualitative, low sensitivity, etc., to achieve the effect of improving sensitivity, small intra-batch variation, and high sensitivity

Inactive Publication Date: 2018-01-16
杭州微瑞科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] There are enzyme-labeled kits for the detection of canine distemper virus neutralizing antibodies on the market, but the ELISA kits require a microplate reader, incubation reaction conditions, plate washing conditions, operating environment and professional technicians, and the detection of samples also requires complex procedures. Pre-treatment, such as collecting blood, separating serum, etc.; and although rapid colloidal gold detection is convenient and does not require professional technicians and working environment, its detection sensitivity is low, and it can only be qualitative, and the detection range is narrow

Method used

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  • Rapid quantitative detection card for canine distemper virus antibody and using method
  • Rapid quantitative detection card for canine distemper virus antibody and using method
  • Rapid quantitative detection card for canine distemper virus antibody and using method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] Embodiment 1, eukaryotic expression of canine distemper virus structural protein H protein

[0034]Step 1. Primer design and PCR amplification of the target gene

[0035] Design specific upstream primer SP1: 5'-TCGAATTCATGAAAAAACTATTTGTG-3' according to the nucleotide sequence of canine distemper virus H gene published in GenBank database; downstream primer SP2: 5'-TTGCGGCCGCTTACATCACATGGCGT-3', the italic black part is the enzyme cut site. Pichia P1: 5'-GACTGGTTCCAATTGACAAGC-3'; P2: 5'-GCAAATGGCATTCTGACATCC-3'. The synthetic hemagglutinin gene (Attachment protein gene, H) genome clone was used as a template, primers SP1 and SP2 were used for PCR amplification reaction, and the final product was checked by 0.8% agarose gel electrophoresis.

[0036] Step 2. Construction and identification of cloning vector TS

[0037] After the target fragment was recovered by the gel recovery kit, it was ligated with the pBS-T vector at 16°C and transformed into Escherichia coli comp...

Embodiment 2

[0048] Example 2: Canine Distemper Virus Structural Protein H Protein Using Specific Synthetic Peptides to Prepare Antigen

[0049] A method for preparing the above-mentioned canine distemper virus structural protein H protein, comprising the following steps:

[0050] Step 1. Sequence analysis of canine distemper virus structural protein H protein: use bioinformatics to predict MHC class I molecules: verify through relevant websites or use computer software to analyze the sequence of CDV-H protein to understand its hydrophilicity, hydrophobicity, Domain accessibility, sequence variability, α-helix, β-turn, antigenicity and other parameters, and then comprehensive analysis and homology modeling methods to predict its tertiary structure, from which the antigenic reactive epitope and amino acid residues are predicted , according to the degree of difficulty of the peptide synthesizer for comprehensive analysis design. Each polypeptide chain contains at least one predicted antigen...

Embodiment 3

[0052] Embodiment three, the detection card of preparation canine distemper virus antibody

[0053] Preparation of canine distemper virus antibody detection standard curve: prepare 6 copies of calibration solution containing canine distemper virus antibody (including canine distemper virus antibody standard), the concentrations are 0, 1 / 1024, 1 / 256, 1 / 64, 1 / 16, 1 / 4 (double dilution of canine distemper virus 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 for canine distemper virus antibody. The standard ...

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Abstract

The invention discloses a rapid quantitative detection card for a canine distemper virus antibody and a using method. The rapid quantitative detection card comprises a detection card shell and a teststrip assembled in the detection card shell. The test strip comprising a plastic base plate with pressure-sensitive adhesive. A sample pad, a marker pad, a nitrocellulose membrane and absorbent paperare sequentially pasted on the base plate. The marker pad is composed of a carrier base layer and a marker, wherein the marker is a membrane formed by spraying the carrier base layer with lanthanide fluorescent detection microspheres and lanthanide fluorescent quality control microspheres. The part, coated with a canine distemper virus H protein antigen, of the nitrocellulose membrane is a detection line. The part, coated with an anti-Chicken IgY antibody, of the nitrocellulose membrane is a quality control line. The marker is fluorescent detection microspheres marked with a canine distemper virus structural protein H protein recombinant antigen and fluorescent quality control microspheres marked with the anti-Chicken IgY antibody. By the adoption of the rapid quantitative detection card,on-site rapid quantitative determination of the canine distemper virus antibody can be achieved, and the practical value and the promotional value are higher.

Description

technical field [0001] The invention relates to a test card and its use, in particular to a canine distemper virus antibody rapid quantitative test card and its use method. Background technique [0002] Canine distemper (Canine Distemper, CD) is caused by canine distemper virus (Canine Distemper virus, CDV) infection. CDV belongs to the Morbillivirus genus of the Paramyxoviridae family. It is a single-stranded, linear, negative-sense RNA virus with an envelope, mostly polymorphic, and the size is between 100-250 nm. CDV consists of six structural proteins: N protein (nucleocapsid protein), P protein (phosphoprotein), M protein (matrix protein), F protein (fusion protein), H protein (hemagglutinin protein) and L protein (large protein), of which N, P and L proteins form nucleocapsid with viral RNA; F protein is one of the glycoproteins on the surface of CDV, and is the main heterotype immune cross-antigen, mediating the gap between its envelope and cell membrane The mutual ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/68
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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