ELISA percolation method for rapidly detecting pathogen antibody, kit for detection, and preparation method of kit

Abstract

The present invention provides an ELISA percolation method for rapidly detecting pathogen antibody, a kit for detection, and a preparation method of the kit. According to the present invention, an antigen-antibody reaction is used to make antigen or second antibody be immobilized on a nitrocellulose membrane, the pathogen IgM (IgG) antibody in a serum sample and the corresponding solid-phase antigen or second antibody on the membrane produce specific binding so as to form a complex when the serum sample passes through the nitrocellulose membrane due to a percolation effect while other unrelated substances are filtered, enzyme-labeled antibody or antigen is added and is combined with the antigen-antibody complex on the membrane during the filtering, and a coloration liquid is added to carry out coloration so as to produce the purple blot conveniently observed by naked eyes; and the time consuming of the detection process is short, the sensitivity is high, the accuracy is strong, the shelf life of the kit can achieve more than or equal to 12 months with the stable dilution buffer solution and the coloration prepared through the method, and the whole detection has characteristics of simple and rapid operation process, no requirement of special equipment, high sensitivity, and high accuracy.

Description

technical field [0001] The invention relates to the technical field of biology, in particular to an enzyme-free diafiltration method for rapid detection of pathogen antibodies, a detection kit, and a preparation method thereof. Background technique [0002] The methods currently used for antibody detection mainly include colloidal gold method, immunochromatographic method, fluorescent PCR method, immunofluorescence method or indirect immunofluorescence method, and enzyme-linked immunospot method. Colloidal gold method is a common labeling technique. It uses colloidal gold as a marker and uses specific antigen-antibody reactions to conduct qualitative and semi-quantitative research on antigens and antibodies. With the advantages of strong stability and low price, it has been widely used in clinical testing and other fields. However, the colloidal gold method can only be used for qualitative or semi-quantitative detection. The accuracy is poor, and it is easy to miss weakly po...

AI Technical Summary

Problems solved by technology

Colloidal gold method is a common labeling technique. It uses colloidal gold as a marker and uses specific antigen-antibody reactions to conduct qualitative and semi-quantitative research on antigens and antibodies. With the advantages of strong stability and low price, it has been widely used in clinical testing and other fields, but the colloidal gold method can only be used for qualitative or semi-quantitative detection, with poor accuracy and easy to miss weak positive samples

Immunochromatography is a rapid diagnostic technique that has emerged abroad in recent years. Its principle is to immobilize specific antibodies on a certain zone of the nitrocellulose membrane, and when one end of the dried nitrocellulose is immersed in the sample (urine) or serum), the sample will move forward along the membrane due to capillary action, and when it moves to the area where the antibody is immobilized, the corresponding antigen in the sample will specifically bind to the antibody. Enzyme staining can make the area display a certain color, so as to achieve specific immunodiagnosis. Although immunochromatography is simple to operate and does not require special equipment, the selection and handling of chromatography materials are more difficult

Fluorescent PCR method is a relatively qualitative and standardized method, which is relatively easy to judge, but the method has poor sensitivity and is prone to false positives; although immunofluorescence method or indirect immunofluorescence method has strong detection specificity, high sensitivity, The speed is fast, but it requires expensive immunofluorescence microscope, the staff needs to have certain experience, and the technical procedures are relatively complicated; the enzyme-linked immunospot method is developed based on the ELISA test, which can detect antibody-secreting cells from the single-cell level , and a cellular immunological detection technique that can detect the amount of secretion. Its principle is to use antibodies to capture cytokines secreted by cultured cells and express them in the form of enzyme-linked immunospot color development. Although the detection sensitivity of enzyme-linked immunospot method is Higher, but its operation is time-consuming and requires strict control of test conditions

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