The fluorescein is labeled on the corresponding antibody and directly reacts with the corresponding antigen. The advantages of the ELISA kit are simple method, high specificity and less non-specific fluorescent staining. The disadvantage is that the sensitivity is low; and each inspection of an antigen requires the preparation of a fluorescent antibody.
Immunofluorescence ELISA kit body technology (fluorescence microscopy technology) After the antigen-antibody reaction, a detection method using fluorescence microscopy to determine the results. After the antigen-antibody reaction, a special instrument is used to measure the fluorescence intensity to estimate the concentration of the analyte.
(1) Fluorescent substances Many substances can produce fluorescence phenomena, but not all ELISA kits can be used as fluorescent pigments. Only those organic compounds that can produce significant fluorescence and can be used as dyes can be called immunofluorescent pigments or fluorescent dyes.
(2) Fluoresceinisothiocyanate (FITC) is yellow or orange-yellow crystalline powder, easily soluble in solvents such as water or alcohol. The molecular weight is 389.4, the maximum absorption light wavelength is 490-495nm, and the maximum emission light wavelength is 520-530nm, showing bright yellow-green fluorescence. The structural formula is as follows: There are two isomeric structures, of which isomer type â… Stability, protein binding ability, etc. are better. It can be stored in a cold and dry place for many years. It is the most widely used fluorescein.
Technical characteristics of immunofluorescence ELISA kit:
The main features of this technology are: strong specificity, high sensitivity and fast speed.
The main disadvantages are: the problem of non-specific staining has not been completely solved, the objectivity of the result judgment is insufficient, and the technical procedures are also relatively complicated. Fluorescence immunoassay can be further divided into several types according to the reaction system and quantitative method. Compared with the radioimmunoassay, the fluorescent immunoassay has no radioactive contamination, and most of them are easy to operate and easy to popularize. A considerable part of the kits for TDM produced abroad belong to this category, and there are also automatic analyzers specially designed for TDM fluorescence polarization immunoassay. ELISA kits have some difficulties in quantitative determination due to the problem of high background in general fluorescence measurement. In recent years, several special fluorescent immunoassays have been developed, which are used in clinical testing as well as enzyme immunoassays and radioimmunoassays.
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