A spectrophotometric assay for nanogram quantities of biotin and avidin.
Parameters and conditions of an enzyme based assay for biotin and avidin are presented. Biotinylated glucose-6-phosphate dehydrogenase when complexed with avidin becomes inactivated. Thus it was possible to construct a competitive assay system for biotin. The assay is sensitive between 100-500 ng/ml and could detect as little as 10 ng in 0.1 ml with a between run error of 2.4%. It requires a 60 min incubation at 21 degrees C and 5 min to assay. The avidin assay, based on the degree of inactivation of biotinylated-glucose-6-phosphate dehydrogenase in relation to the concentration of avidin, could detect as little as 0.25 ng in 0.1 ml or 2.5 ng/ml with an assay time of 10 min with a between run error of 3.9%. Both assays are rapid with significant improvements over other non-isotopic methods in sensitivity and comparable to radioisotopic methods in sensitivity with the added advantage of ease of method.
D001360:Avidin / Q000032:analysis*; D001710:Biotin / Q000032:analysis*; D005954:Glucosephosphate Dehydrogenase; D008832:Microchemistry / Q000379:methods; D013053:Spectrophotometry / Q000379:methods*
D001360:Avidin; D001710:Biotin; D005954:Glucosephosphate Dehydrogenase