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Recombinant Protein Purification using Strep-Tactin®XT

IBA’s 3rd generation Strep-tag® system is based on the Strep-Tactin®XT resin in combination with Twin-Strep-tag® and is one of the most widely used purification systems for recombinant proteins. Due to a binding affinity of Strep-Tactin®XT to Twin-Strep-tag® in the low pM range (near covalent binding!), the system is superior to all other available affinity purification systems and can be used for a wide variety of applications, e.g. protein purification and assay development.

Development of the Strep-tag® system
Development of the Strep-tag® system


Strep-Tactin®XT provides superior performance in the field of high throughput screening, batch purification, purification using denaturing conditions and protein interaction studies. Furthermore higher protein yields compared to Strep-Tactin® are obtained. Simultaneously, Strep-Tactin®XT preserves all benefits of Strep-Tactin® for protein purification, e.g. high purity (> 95 %), physiological buffer conditions, mild elution, regeneration of the resin for re-use.

Get highly pure proteins with Strep-Tactin®XT resin

One of the key advantageous of the Strep-tag® system is to achieve unparalleled protein purity (> 95 %) after purification of recombinant proteins independent of the expression host. This makes the system advantegous to other affinity purification systems like His-tag.

The 3rd generation Strep-tag® system with its new features makes it suitable for additional applications:

New applications: Excellent performance:
  • High-throughput screening
  • Batch purification
  • Denaturating conditions
  • High protein yields and purity
  • Optimized elution profile for highly concentrated target protein

When should the Strep-tag® system be the method of choice?

Besides the purification of classical proteins the Strep-tag® system should be the method of choice for:

  • metalloproteins
  • membrane proteins
  • low abundant proteins (in combination with Strep-Tactin®XT)
  • sensitive protein complexes with multiple subunits
  • bioactive proteins
  • and any other protein
Strep-Tactin®XT:Twin-Strep-tag® share a near covalent binding affinity
Comparison of ligands and receptors within the Strep-tag® system regarding binding affinities
Comparison of ligands and receptors within the Strep-tag® system regarding binding affinities


The Streptavidin:Biotin binding is the strongest noncovalent biological interaction known. With IBA's newly developed Strep-Tactin®XT in combination with Twin-Strep-tag®, this high binding affinity can be used for e.g. protein purification, protein interaction analysis or assays (e.g. BIAcore). Since Biotin is still binding strongly to Strep-Tactin®XT it can be used for elution of Twin-Strep-tag® fusion proteins.

Strep-Tactin®XT purification cycle
Strep-Tactin®XT purification cycle
Strep-Tactin®XT purification cycle

Purification of target proteins in a simple one-step purification cycle:

Step 1: The cell lysate / culture supernatant is applied on the column.

Step 2: Once the tagged protein has bound specifically to Strep-Tactin®XT the host proteins are instantly washed away with moderate amounts of physiological wash buffer (Buffer W).

Step 3: Bound Twin-Strep-tag® protein is gently eluted by Buffer BXT (wash buffer containing 50mM biotin) which specifically competes for the biotin binding pocket.

Step 4: Regeneration of the column is achieved by the application of 3 M MgCl2. Magnesium chloride removes the biotin from the binding pocket. MgClcan be removed simply by applying Buffer W. Instead of MgCl2, freshly prepared 10 mM NaOH can also be used. Strep-Tactin®XT resin can be regenerated and re-used 3 to 5 times without loss in performance.

Strep-tag®II and Twin-Strep-tag®

Strep-tactin®XT can be used in combination with the following tags:


  • 8 amino acids
  • sequence: Trp-Ser-His-Pro-Gln-Phe-Glu-Lys (WSHPQFEK)


  • Two Strep-tag®II motifs in series
  • Higher affinity than Strep-tag®II

Features of both tags

  • Fusion to the N- or C-terminus
  • No influence on protein folding and function
  • No removal necessary