Method for detecting SPA FC fragment bound mammalian blood serum total IgG by dolloidal gold marker protein A

Abstract

A method for detecting total IgG combined with Fc fragment of Staphylococci protein A (SPA) in mammalian serum with colloidal gold-labeled protein A comprises the following steps of: 1) construction of a standard curve and calculation of regression equation for each batch of colloidal gold-labeled probe and micro-reaction by (1) setting 8 standard holes, adding 50MuL 0.01M TBS solution (pH 7.4, containing 0.1% calf serum) to each hole; (2) adding 50MuL standard IgG solution from the first hole to the sixth hole by doubling dilution, sucking out 50MuL TBS solution from the seventh hole, adding 50MuL standard IgG solution, and leaving the eighth hole as a blank control; (3) adding 50MuL colloidal gold-labeled probe diluted by 2-6 times in the reaction holes; (4) completely mixing the reaction plate, and reacting at room temperature for 20-40min; (5) detecting absorbance at 620nm with a microplate-reader for ELISA; and (6) calculating common logarithm of the absorbance associated with the standard IgG; and 2) sample detection by (1) adding 50MuL sample solution in the sample holes; (2) adding 50MuL diluted colloidal gold-labeled probe in the sample holes; (3) completely mixing the reaction plate, and reacting at room temperature for 20-40min; and (4) detecting absorbance of each hole at 620nm with the microplate-reader for ELISA, and calculating corresponding IgG values according to the regression equation of the standard curve.

Description

technical field [0001] The invention belongs to the field of detection reagents, in particular to a method for detecting the total amount of IgG in mammalian serum combined with FC fragments of SPA by colloidal gold-labeled protein A. Background technique [0002] Immunoglobulin (IgG) is a protein widely distributed in animal cerebrospinal fluid and serum, which can promote the phagocytosis of pathogens by immune cells; promote the killing and destruction of tumor cells or infected cells by immune cells; It can agglutinate with the pathogen when it comes into contact with it; it is the only immunoglobulin that can be passed to the fetus through the placenta, which can enhance the immunity of the fetus and newborn. Detecting its content is of great significance for understanding the immune status and health of animals. [0003] There are many methods for detecting the total amount of IgG in serum, such as: one-way immunodiffusion method, enzyme-linked immunosorbent assay (EL...

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Problems solved by technology

[0003] There are many existing methods for detecting the total amount of IgG in serum, such as: one-way immunodiffusion method and enzyme-linked immunosorbent assay (ELISA), etc. are currently the two most widely used methods, and one-way immunodiffusion method is the most classic method. This method is characterized by simple operation, simple equipment, and low cost of determination. Therefore, this method has been adopted internationally for many years, but its accuracy and sensitivity are not very good.

In addition, there is enzyme-linked immunosorbent assay, although the accuracy and sensitivity of this method are good, but the operation is complicated, the equipment is expensive, and the cost is high, so it is only used in some researches.

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