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Recombinant Protein Purification using Strep-Tactin®XT

IBA’s 3rd generation Strep-tag® system is based on the novel Strep-Tactin®XT Superflow® 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 Superflow 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 10mM NaOH. Sodium hydroxide removes the biotin from the binding pocket. NaOH can be removed simply by applying Buffer W. 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