Indirect ELISA detection method of SARS-CoV-2S protein IgM

Abstract

The invention relates to an indirect ELISA (Enzyme-Linked Immuno Sorbent Assay) detection method of SARS-CoV-2S protein IgM (Immunoglobulin M); the method comprises the following steps: adding new coronavirus spike protein diluted by adopting an ELISA coating solution into an elisa plate with the standard of 100 micro-mililiter per hole, staying overnight at 4 DEG C, and performing cleaning with a PBST (Phosphate Buffer Solution); adding 250 microlitres of a 5% skim milk powder solution for sealing, diluting the serum to be detected with an ELISA antibody diluent as a primary antibody, adding 100 microlitres of the primary antibody into each hole, and performing incubating at 37 DEG C for 1 h; and using horseradish peroxide labeled anti-human IgM diluted by an ELISA antibody diluent as a secondary antibody, adding 100 microlitre of the secondary antibody into each hole, performing incubation at 37 DEG C for 45 min, adding 100 microlitre of a single-component TMB color developing solution into each hole in a dark place, performing incubation at 37 DEG C for 5 min, adding 50 microlitres of a stop solution into each hole under the conditions of setting the detection wavelength to be 450 nm, the reference wavelength to be 630 nm, and the absorbance value OD450 nm detected by a microplate reader to be 450 nm.

Description

technical field [0001] The invention belongs to the technical field of virus detection, and relates to an indirect ELISA detection method of SARS-CoV-2 S protein IgM. Background technique [0002] The new coronavirus (2019-nCoV), currently known as severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2), causes coronavirus disease 2019 (COVID-19). SARS-CoV-2 has a high degree of homology with another human coronavirus (severe acute respiratory syndrome coronavirus (SARS-CoV)). SARS-CoV-2 is transmitted by droplets and possibly by the fecal-oral route. The symptoms of COVID-19 cases are dry cough, fever, body aches and weakness, and diarrhea. The clinical manifestations of severe cases are high fever, dyspnea, and low blood oxygen levels. Most of the deaths are caused by respiratory failure and multi-organ damage caused by cytokine storms. Exhausted. The homotrimer on the surface of the coronavirus mediates the entry of the virus into the host, which is a transme...

AI Technical Summary

Problems solved by technology

At present, several polymerase chain reaction (PCR)-based technologies to detect viral RNA are the main methods to determine the diagnosis of new coronavirus infection, but the positive rate of nucleic acid detection based on respiratory specimens is low, and a single nucleic acid test result may cause missed diagnosis of new coronavirus patients. Other methods need to be developed to supplement nucleic acid detection. At the same time, real-time fluorescent quantitative PCR (quantitative real-time PCR RT-qPCR) can only detect the presence of virus in the specimen, but it cannot reflect the humoral immune response of the case

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