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Kit supplied with the following:
By comparing a known amount of a 100% biotinylated AviTagTM fusion protein as a standard against a known amount of BirA-reacted protein-of-interest in an ELISA-type, multi-well dilution format (called BRTA for Biotinylation Reaction Titration Assay) it is possible to assess the extent of biotinylation achieved in your own BirA biotinylation reactions in a highly accurate and reproducible manner.
In addition, a mass shift between biotinylated and unbiotinylated fusion proteins visible on both SDS-PAGE and Western blot can be used to subjectively determine the amount of biotinylation that has occurred in a reaction by the observation of differences in band density in a mixed population of biotinylated vs. nonbiotinylated substrate.
The BIS-300 Kit contains the fully biotinylated standard used for the BRTA protocol and an unbiotinylated AviTag"d protein to react in a parallel to your BirA/protein biotinylation reaction as a positive control. A link to the BRTA protocol can be found here.
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