BioLock - Biotin Blocking Solution
Biotin is often present in cell culture media. Its nearly irreversible binding to Strep-Tactin® has to be taken into account when recombinant proteins, secreted to the cell culture supernatant, are aimed to be directly purified via Strep-Tactin® affinity chromatography for two reasons:
- It efficiently competes with the (Twin-)Strep-tag® target protein, thereby preventing it from binding to the resin
- it inactivates Strep-Tactin® resins and thus, makes re-use impossible.
BioLock is a ready-to-use and cost effective
solution for the removal of free biotin from cell culture supernatant.
It simply has to be added to the cell culture supernatant prior to
purification.
The required amount of BioLock for effective biotin blocking can
easily be determined since 1 ml BioLock blocks 70 μg biotin. In addition
we provide a table with the required BioLock amounts for standard
mammalian and insect cell culture media:
Biotin Blocking
For blocking biotinylated proteins in Western blots (like BCCP protein from E. coli), the membrane is incubated with Biotin Blocking Buffer. Use a dilution of 1:1000 in standard Western blot blocking reagent prior to detecting Strep-tag® proteins with Strep-Tactin® conjugates.
Avidin prevents biotin and biotinylated proteins from binding to Strep-Tactin® columns during Strep-tag® protein purification. This is important if growth media containing free Biotin are used (for further information see “Biotin Blocking”).
