Fluorescence lateral chromatography detection method for mouse poxvirus antibody

The fluorescence lateral flow assay method for mousepox virus detection addresses interference and cross-reactivity issues in existing assays by using PCR-amplified A29L gene and recombinant proteins, achieving high sensitivity and specificity, and ensuring accurate detection of mousepox virus antibodies in clinical samples.

CN119716046APending Publication Date: 2025-03-28GUANGDONG BIOTECHNOLOGY RESEARCH INSTITUTE (GUANGDONG PROVINCE EXPERIMENTAL ANIMAL MONITORING CENTER)
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Patent Information

Application Number
CN202411647649.2
Authority / Receiving Office
CN · China
Patent Type
Applications(China)
Current Assignee / Owner
Filing Date
2024-11-18
Publication Date
2025-03-28

AI Technical Summary

Technical Problem

Current immunodot assays for detecting mousepox virus (ECTV) are prone to interference from environmental and sample quantity factors, and can produce false positives and negatives, with potential cross-reactivity issues.

Method used

A fluorescence lateral flow assay method involving PCR amplification of the A29L gene, construction of a recombinant protein, and formation of fluorescent probes to enhance specificity and sensitivity, using a designed immunochromatographic test strip with a combination of BSA, NaCl, PEG4000, Triton X-100, and borate buffer.

Benefits of technology

The method achieves high sensitivity and specificity, with 100% concordance in clinical samples compared to ELISA, maintaining accuracy even at high dilutions, and is simpler, faster, and more cost-effective, effectively identifying low concentrations of mousepox virus antibodies.

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Abstract

The invention relates to the technical field of biomedicine, and discloses a mouse poxvirus antibody fluorescent lateral chromatography detection method which comprises the following steps: step 1, extracting nucleic acid of mouse poxvirus ECT, and amplifying an A29L gene by using a PCR technology to obtain an amplification product of the gene of 333bp; step 2, connecting the amplification product with a pGEX-6P-1 vector through double enzyme digestion, and constructing a recombinant plasmid; step 3, transforming the recombinant plasmid into escherichia coli E.coli DH5alpha for expression, so as to obtain a recombinant protein; the preparation method comprises the following steps of 1, preparing a recombinant protein, 2, preparing a mouse poxvirus antibody, 3, preparing a mouse poxvirus antibody, 4, verifying the immune characteristic and specificity of the recombinant protein through Western Blot and ELISA methods, and 5, preparing fluorescent microspheres and coupling the fluorescent microspheres with the mouse poxvirus antibody to form a fluorescently-labeled antibody probe. Compared with the traditional rapid detection technology, the kit has the advantages of good sensitivity and specificity, reduction of the influence of interfering substances in a sample, accurate and reliable detection result, wide detection range, simple and rapid operation and low cost.
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