Biotin Pincer^TMAssay

Catalog No: BT1006

Biotin, also known as vitamin H or B7, is a water-soluble B-complex vitamin which plays an indispensable role in numerous naturally occurring carboxylation reactions in living cells^1,2. Biotin is also very commonly used to label peptides, DNA or proteins because it is a relatively small molecule and can be conjugated to many biological molecules without altering the biological activity of the molecules. The strong binding between biotin and the tetrameric protein avidin, streptavidin, or neutravidin (with a dissociation constant Kd in the order of 10-14 M)³ allow biotin to be used in different biotechnological applications. The Biotin PincerTM Assay Kit provides a convenient method to quantify the biotin concentration in unknown solution as well as estimate the amount of biotin in biotinylated samples.

This assay consist two biotin modified oligonucleotides (with short complementary sequences) labeled with fluorescent probes (pincers) and streptavidin protein⁴. The pincers will form a complex with streptavidin and have increased FRET (Fluorescence resonance energy transfer).   When biotin is present, it will compete with the biotinylated pincer and result in decreased FRET.  The amount of FRET signal is associated with the biotin concentration presented in the samples. This assay is easy to perform and the whole procedure can be completed within 40 minutes.

Biotin Pincer^TM Assay - Standard Curves

The following biotin standard curves were prepared using the Biotin Pincer^TM Assay with the fluorescence signal read after 30 minuets after addition of Biotin Pincer^TM Assay Solution B. The standard errors were from triplicate measurements.

Biotin Pincer^TM Assay – Sensitivity

The minimum detectable dose of biotin using as a standard curve is 2 nM.

Reference:

1. Moss J, Lane MD., (1971) The biotin-dependent enzymes. Adv Enzymol. 35, 321-442.
2. Wakil, SJ., Titchener, EB., & Gibson, DM., (1958) Evidence for the participation of biotin in the enzymic synthesis of fatty acids. Biochem. Biophys. Acta. 29, 225-226.
3. Green, NM., (1990) Avidin and streptavidin. Methods Enzymol. 184, 51-67.
4. Heyduk, T, Tian, L. Molecular Biosensors for Use in Competition Assays. US Patent Application 11/836,339.

RESEARCH USE:

For research use only, not for use in diagnostic procedures.

Patent pending.

Product name	Measurements	Cat. #	Protocol	MSDS	Price	Order
Biotin PincerTMAssay kit	100	BT1006-100			$150
Biotin PincerTMAssay kit	384	BT1006-384			$320
384-well round-bottom low volume non-binding surface black polystyrene microplate	1 plate	163301	--		$9.50