Buffers & Reagents for Protein Purification with Strep-Tactin®

For removal of unbound proteins and other material during Strep-tag® protein purification via Strep-Tactin^® and Strep-Tactin^®XT columns.

Desthiobiotin displaces Strep-tag^®II proteins at the biotin-binding site of Strep-Tactin^® in a competitive manner resulting in a mild elution of the protein. For your convenience, we offer desthiobiotin ready-to-use in solution (Buffer E). If you wish to prepare your own elution buffer, desthiobiotin is also offered as a lyophilized powder. Please note that after elution with Buffer E, the Strep-Tactin^® column can be regenerated for atleast 3-5 times with HABA.

Do not use in combination with Strep-Tactin^®XT!!!

Contains HABA (2-[4'-hydroxy-benzeneazo]benzoic acid), which displaces desthiobiotin at the biotin-binding site regenerating Strep-Tactin®. The yellow HABA solution turns red when desthiobiotin has been removed from Strep-Tactin®, clearly indicating the regeneration process and activity status of the column.

Do not use in combination with Strep-Tactin^®XT!!!

Convenient set for the purification of recombinant Strep-tag^® fusion proteins on immobilized Strep-Tactin^® and for the regeneration of the columns. Consists of Buffers W / E / R.

Do not use in combination with Strep-Tactin^®XT!!!

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 1ml BioLock blocks 70 μg biotin. In addition we provide a table with the required BioLock amounts for standard mammalian and insect cell culture media at
www.iba-lifesciences.com/technical-support.html >> troubleshooting>> Biotin Blocking

Elution buffer for competitive displacement of purified recombinant Strep-tag^® or Twin-Strep-tag^® fusion proteins from immobilized Strep-Tactin^®. The buffer is recommended for the elution from Strep-Tactin^® Spin columns.
This buffer is also recommended for use with Twin-Strep-tag^® in protein complex or PPI applications, since it contains Biotin, which has a higher affinity to Strep-Tactin^® and therefore, more efficiently competes with the (Twin)Strep-tag^® binding. The regeneration of the Strep-Tactin^® columns in PPI applications is not recommended in order to avoid cross contaminations.

Please note that after elution with Buffer BE the resin cannot be regenerated.

Do not use in combination with Strep-Tactin^®XT!!!

Is used for induction of tet promoter controlled pASK-IBA vectors. The E. coli expression cassette of the Strep-tag®/Strep-Tactin® overexpression system is under transcriptional control of the tetA promoter/operator and repressor. The promoter is induced by a low concentration of anhydrotetracycline (AHT) saving costs and minimizing the antibacterial influence of AHT.

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”).

For blocking biotinylated proteins in Western blots (like BCCP protein from E. coli, see Applications), 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.

For specific customer purposes.