Competitive inhibition enzyme-linked immunosorbent assay (ELISA) method for grass carp tumor necrosis factors alpha

Abstract

The invention relates to a competitive inhibition enzyme-linked immunosorbent assay (ELISA) method for grass carp tumor necrosis factors alpha. The method comprises the following steps of: 1, determining the concentration of coating antigens and the optimal dilution ratio of polyclonal antibodies; 2, establishing the competitive inhibition ELISA method, and coating an ELISA plate of grass carp tumor necrosis factor alpha protein to obtain antigens immobilized on the ELISA plate; mixing enzyme-labeled antibodies and an antigen mixed solution, and performing competitive binding on the solution and the antigens immobilized on the ELISA plate; washing the ELISA plate and developing the color of the substrate; and reading holes of the ELISA plate to obtain the light absorption value of each hole, calculating the inhibition rates of a standard antigen and a sample to be detected according to the read light absorption values respectively, and calculating the concentration of the sample to be detected according to the inhibition rates. The method has the advantages that a quick and efficient detection means is provided for detecting the protein expression level of the grass carp tumor necrosis factors alpha; and the method is stable, reliable and low in cost.

Description

technical field [0001] The invention belongs to the technical field of biological detection, in particular to the field of enzyme-linked immunoassay (ELISA) biological technology. Background technique [0002] Tumor necrosis factor alpha (tumor necrosis factor-alpha, TNF-α) is an important cytokine. As early as more than 30 years ago, Carswell et al. discovered a protein substance present in serum that can inhibit and kill certain tumor cells and cause hemorrhage and necrosis of tumors in vivo, which was named tumor necrosis factor (TNF). ). Studies have found that tumor necrosis factor alpha participates in and regulates the body's immune function and inflammatory response, and plays an important role in promoting cell growth, differentiation, and apoptosis. Since the first full-length cDNA sequence of tumor necrosis factor alpha gene was cloned from flounder, the research on tumor necrosis factor alpha in fish has been paid attention to, and the research shows that fish ...

AI Technical Summary

Problems solved by technology

At the same time, due to the large gap in structure and function between fish TNF-alpha and mammalian and amphibian TNF-alpha molecules, TNF-alpha proteins and antibodies from other races cannot be used to replace fish TNF-alpha research, coupled with the lack of antibodies to TNF-alpha in fish, has bottlenecked research on the functional status of TNF-alpha

At present, there is no specific enzyme-linked immunoassay method for detecting the concentration of TNF-alpha in grass carp

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