Introduction
Protein Research Applications
Biotin and peptide binding via amide bond
Biotin-labeled peptides are commonly detected or purified using avidin conjugates for many protein-based research applications, including western blotting, ELISA, immunoprecipitation (IP), cell surface labeling, affinity purification, immunohistochemistry (IHC), and flow cytometry/fluorescence-activated cell sorting (FACS).
Protein interaction complexes, including those involving surface proteins, can be determined using a specific biotinylated peptide as the bait to capture potential binding partners from a biological sample. An example of such a protocol uses streptavidin to capture biotinylated proteins in vivo in a single step, followed by and tandem affinity purification using a biotinylated peptide coupled to a traditional affinity tag (such as FLAG or 6×His). The biotinylated peptides are immobilized to an affinity support material such as avidin agarose:
Incubate the crude samples with the solid support
Remove unbound sample components
Elute the target molecule that is bound to its interacting proteins by altering the buffer used for elution.