KRIBIOLISA™ Darbepoetin (ARANESP™) ELISA
Enzyme Immunoassay for the Quantitative Determination of Darbepoetin in human serum and plasma
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Introduction:
Erythropoietin (EPO) is a heavily glycosylated protein with a molecular weight of about 30,000 - 34,000 Daltons. Human EPO is a polypeptide consisting of 165 amino acids, containing one O-linked and three Nlinked carbohydrate chains. The recombinant EPO is a good substitute for the native protein for use in an immunoassay. Darbepoetin is a re-engineered form of erythropoietin, contains 5 N-linked oligosaccharide chains and has a molecular weight of 37,100 Daltons and a carbohydrate composition of 51%.It has a 3-fold longer serum half-life compared to epoetin alpha and epoetin beta.
Intended Use:
The KRIBIOLISA™ Darbepoetin (ARANESP) ELISA is used as an analytical tool for quantitative determination of Darbepoetin in human serum and plasma.
Principle:
The method employs the quantitative sandwich enzyme immunoassay technique Standards or samples are pipetted into microwells pre-coated with Anti-Darbepoetin antibody and Darbepoetin present in the sample and standards are bound by Anti-Darbepoetin antibody. In the second step, Detection antidody is added and incubated. In the third step, a HRP conjugate is pipetted and incubated. Free HRP conjugate will be removed by washing. Addition of TMB substrate will develop blue color and intensity of blue colour in wells is proportional to the concentration of Darbepoetin present in standard or sample. Color development is then stopped by addition of stop solution. Absorbance is measured at 450 nm.
