Method for detecting protein content

Abstract

The invention relates to a method for detecting protein content. The method comprises the following steps of: preparing standard protein samples and reagents (1), (2) and (3); preparing the standard protein samples with gradient; adding the reagents (1), (2) and (3) into the gradient samples and samples to be tested by a micropore method or a standard test tube method; measuring absorbance; drawing a standard curve; and calculating the concentration according to the absorbance of the samples to be tested. By the method, the protein content can be detected accurately. The method is applicable to protein solution containing a plurality of interfering substances. By the method, the inclusiveness for the interfering substances such as a surfactant, a reducing agent, a chelating agent, a nitrogen-containing compound and the like in the protein analysis and detection process can be improved. The method is a simple, quick, high-sensitivity and high-stability method for detecting the protein concentration and has a wide application prospect in the field of life science research.

Description

technical field [0001] The invention belongs to the technical field of biochemical analysis and detection, and in particular relates to a protein content detection method. Background technique [0002] Conventional protein concentration detection methods mainly include Lowry method, BCA (Bicinchoninic acid) method and Bradford method. The Lowry method is mainly based on the Cu under alkaline conditions 2+ Form a blue complex with the -CO-NH- group of the peptide backbone, and then reduce the phosphotungstic acid and phosphomolybdic acid in the Folin's phenol reagent to tungsten and molybdenum blue for further color development. The BCA method is to use BCA to react with the Cu formed in the first step of the Lowry method. + It is measured by the blue-purple complex formed by binding. The basis of the Bradford method is that the dye Coomassie Brilliant Blue G-250 can combine with lysine and arginine in proteins or peptides with a molecular weight above 3 kD to form colored...

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Problems solved by technology

[0003]The Lowry method is cumbersome to operate and is also affected by many chemical components, such as when the content of surfactant Triton X-100 or SDS is only 1%, or metal ions When the concentration of chelating agent EDTA reaches 10 mM, or the content of urea reaches 3 M, or the concentration of ammonium sulfate reaches 0.25 M, etc., it will seriously interfere with the detection results; although the BCA method is less affected by the interference of surfactants, it is not affected by reducing agents and metal ion chelation. Sensitive to the presence of mixture and ammonium sulfate

The Bradford method is simple to operate, but similar to the Lowry method, it has poor tolerance to surfactants. When 0.1% SDS or 0.5% TritonX-100 exists, the absorbance of protein samples will be greatly affected

Therefore, the specific use of these methods will always be limited due to the composition of the assay solution or protein lysate. In other words, if there are interfering substances, different methods must be adopted for different interfering substances in the determination of protein concentration. Inconvenience

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