Dipstick based immunochemiluminescence biosensor for the analysis of vitamin B12 in energy drinks: A novel approach

In this article, we describe a dipstick based immunochemiluminescence (immuno-CL) biosensor for the detection of vitamin B-12 in energy drinks. The method is a direct competitive type format involving the immobilization of vitamin B-12 antibody on nitrocellulose membrane (NC) followed by treatment with vitamin B-12 and vitamin B-12-alkaline phosphatase conjugate to facilitate the competitive binding. The dipstick was further treated with substrate disodium 2-chloro-5-(4-methoxyspiro {1,2-dioxetane-3,2(sic)-(5(sic)-chloro)tricyclo[3.3.1.13.7]decan}-1-4-y1)-1-phenyl phosphate (CDP-Star) to generate chemiluminescence (CL). The number of photons generated was inversely proportional to the vitamin B-12 concentration. After systematic optimization, the limit of detection was 1 ng mL(-1). The coefficient of variation was below 0.2% for both intra- and inter-assay precision. Vitamin B-12 was extracted from energy drinks with recovery ranged from 90 to 99.4%. Two different energy drinks samples were analyzed, and a good correlation was observed when the data were compared with a reference enzyme linked immuno sorbent assay (ELISA) method. The developed method is suitable for an accurate, sensitive, and high-throughput screening of vitamin B-12 in energy drinks samples. The dipstick technique based on immuno-CL is suitable for the detection of several analyte in food and environmental samples. (c) 2012 Elsevier B.V. All rights reserved.