pleased to announce the release of the FRET based biotin quantitation kit.
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.
- Angerer, L. et al. Cell.
9, 81-90 (1976).
- Gitlin,G. et al. Biochem. J.