Biotin/avidin complexes are widely applied for a variety of biological detections. Although a large number of biotin-labeled bioconjugates are commercially available, the accurate determination of biotinylation degree (ratio of biotin/biopolymer) is still a great challenge for biochemists. HABA is predominantly used for determining the degree of biotinylation (through its absorption with the extinction coefficient = 34,000/M-1cm-1). When a biotin-containing sample is added, the biotin binds strongly to avidin and displaces the weakly bound HABA. The resulting decrease in absorbance relates to the amount of biotin. However there are many factors that affect the accuracy of HABA method, making this method unreliable for many biotin-labeled conjugates.
Fig. 1
The Chemical structure of ReadiView™ biotin. RM = reactive moiety for labeling purpose; CT = color tag for calculating biotinylation degree.
Our ReadiView™ biotin contains specially designed Color Tag (CT) that makes the biotinylation degree readily accessible by simply calculating the corrected absorption ratio of 280 nm/385 nm. Our specially designed tag has a very minimal effect on the biotin binding affinity, and its absorption maximum is designed to make the tag have minimal quenching effect on most fluorophores used for labeling avidins.