AnaSpec is pleased to announce the release of the FRET based biotin quantitation kit. The SensoLyte^® Biotin Quantitation Assay Kit provides a convenient way to estimate the efficiency of biotinylation for biotin-protein conjugates. The kit also can be used to quantitate biotin concentration in a solution.

In the first step, Reagent A, which is tagged with a quencher dye, is briefly incubated with biotin samples/standards. In the second step, fluorescent biotin, Reagent B, is added to assay mixture. The assay can detect between 2 to 20 picomoles of biotin in a sample (Fig. 1). A biotinylated IgG for use as a positive control and a protease for optional protein digestion to expose hidden biotin groups are also included in the kit.

Figure 1. Biocytin standard calibration curve. Serial dilutions of Biocytin were mixed with Reagent A, followed by Reagent B. After 5 minute incubation, fluorescence was measured at Ex/Em=490/520 nm (FlexStation 384II).

Biotin and avidin (or streptavidin) bind non-covalently with a higher binding affinity than most antigen-antibody interactions.^{1, 2} This very tight binding makes labeling proteins with biotin a useful tool for applications such as affinity chromatography and immunoanalytical methods. Reaction conditions for biotinylation are chosen such that the target molecule (e.g. an antibody) is labeled with sufficient biotin residues to purify or detect the molecule, but not too much that the biotin will interfere with the function of the molecule.