Download the Strep-tag II purification cycle showing a good overview of the convenient procedure (PDF, 95 kb)
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Features and benefits
- Simple one-step purification from crude lysate to 99% pure protein under physiological conditions possible (for example click here)
- High and selective binding affinity and high capacity due to special Strep-Tactin resins
- Resins for gravity flow, low pressure or HPLC applications
- Co-purification of non-covalently bound ligands, thus, protein-protein interaction studies are possible
- Column regeneration and activity status is visualized by color change
Principle and properties
The Strep-tag purification system is based on the highly selective binding of engineered streptavidin, called Strep-Tactin, to Strep-tag II fusion proteins. This technology allows one-step purification of almost any recombinant protein under physiological conditions, thus preserving its bioactivity. The Strep-tag system can be used to purify Strep-tag II proteins from any expression system including baculovirus, mammalian cells, yeast, and bacteria. For vectors for bacterial expression click here.
Procedure
The purification of Strep-tag II fusion proteins is very simple and user-friendly. The complete procedure can be performed under nearly physiological conditions, e.g. in PBS buffer and 2.5 mM desthiobiotin in PBS buffer during elution.
- Once the tagged protein has bound specifically to the column the unspecific proteins are rapidly washed away with small amounts of physiological washing buffer.
- Then, small amounts of the specific competitor desthiobiotin are added in order to elute the Strep-tag protein. Since the buffer conditions during elution essentially remain unchanged, potentially unspecifically binding proteins (without Strep-tag) will not be eluted and, thus, will not contaminate the protein of interest.
- To regenerate the column desthiobiotin is displaced by adding the yellow solution HABA (hydroxyazophenyl benzoic acid formula). After the removal of desthiobiotin the column turns red due to the binding of HABA as red colored hydrazone isomer to Strep-Tactin. This clear color change indicates the regeneration and activity status of the column.
- HABA can be removed simply by using washing buffer. Once the red color has disappeared the column can be re-used.
The Strep-tag II purification cycle gives you an illustrated overview of the procedure.
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