A rapid detection kit for novel coronavirus antibody based on mixed antigen
A coronavirus, mixed antigen technology, applied in the direction of viruses/phages, viruses, viral peptides, etc., can solve the problem of no specific treatment methods for diseases, and achieve the effects of effective control, high detection sensitivity, and simple operation.
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Embodiment 1
[0037] This embodiment provides a novel coronavirus rapid diagnostic kit, specifically prepared through the following steps:
[0038] 1. Expression and purification of COVID-19 nucleocapsid protein
[0039] 1-1. With the nucleotide sequence corresponding to the amino acid sequence of the COVID-19 nucleocapsid protein (i.e. N protein) as a template, design a forward primer whose nucleotide sequence is ATGTCTGATAATGGACCCCA, and a reverse primer whose nucleotide sequence is GGCCTGAGTTGAGTCAGC Direction primers; PCR amplification, double digestion of PCR products and vectors, and cloning into prokaryotic expression vectors, including but not limited to pET28a / pET22b / pET21b / pET32a / pET40 / pGex-6P-1 / pMal-c2x.
[0040]1-2. Transform the recombinant plasmid containing the nucleotide sequence of the COVID-19 nucleocapsid protein into E.coli BL21 or Rossetta strain, spread it on a plate containing ampicillin or kanamycin, and incubate at 37°C After 16 hours, pick a single colony and inoc...
Embodiment 2
[0055] The COVID-19 nucleocapsid protein antigen colloidal gold and the COVID-19 spike protein antigen colloidal gold prepared by the method in Example 1 were clinically verified.
[0056] Take 5 clinically confirmed cases of novel coronavirus pneumonia, 5 normal samples and 2 negative control samples for verification. The verification method is as follows:
[0057] The COVID-19 nucleocapsid protein antigen colloidal gold and spike protein antigen colloidal gold were respectively coated into 96-well plates, and the coating amount of each well was 0.2 μg / well. In addition, the COVID-19 nucleocapsid protein antigen colloidal gold and COVID-19 spike protein antigen colloidal gold were mixed and coated into a 96-well plate, and the COVID-19 nucleocapsid protein antigen colloidal gold and COVID-19 spike protein antigen Each antigen colloid was coated with 0.2 μg / well. Negative control wells were set up when coating.
[0058] Add 100 μL of the above-mentioned different types of sp...
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