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Strep-Tactin® vs Strep-Tactin®XT

Get all information in an overview:

Strep-Tactin® vs Strep-Tactin®XT

Choose the right resin according to your application:

When to use which resin?

Application Strep-Tactin® Strep-Tactin®XT
  Strep-tag®
Twin-Strep-
tag®
Strep-tag®
Twin-Strep-
tag®
purification
 

 
  • batch
  • gravity flow
  • Äkta purification
  • denaturing conditions
  • low-abundant proteins
  • membrane proteins
detection
assay

What are the differences?

Differences in binding affinities

The Streptavidin:Biotin binding is the strongest non-covalent biological interaction known. With IBA's newly developed Strep-Tactin®XT in combination with Twin-Strep-tag®, this near covalent 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.

Differences in the purification procedure
Click to enlarge the figure

The overall purification procedure of Strep-Tactin®XT and Strep-Tactin® is the same (see figure). Only two changes were introduced:

  • Elution from Strep-Tactin®XT needs to be performed with biotin
  • Regeneration of Strep-Tactin®XT resin will be achieved with 10 mM NaOH
Technical comparison of Strep-Tactin® and Strep-Tactin®XT
Features Strep-Tactin® Strep-Tactin®XT
Buffer compatibility/variability high high
Detergent stability moderate high
Wash stability moderate high
Affinity (tag/ligand)
µM for Strep-tag®
nM for Twin-Strep-tag®
nM for Strep-tag®
pM for Twin-Strep-tag®
pH range 7.5 - 9 5 - 10
Denaturing conditions no yes (up to 6 M urea)
Ease of use yes yes
Elution conditions

desthiobiotin (reversible)

biotin (non-reversible)

biotin (reversible)
Re-use possible yes yes
Regeneration condition HABA (Buffer R) NaOH

Benefits of Strep-Tactin®XT

Strep-Tactin®XT allows higher protein yields

Strep-Tactin®XT allows the purification of higher protein yields compared to Strep-Tactin®. This was determined by purification of different target protein on either Strep-Tactin® or Strep-Tactin®XT. The obtained protein yields were determined and compared in the graph below. Here, we found that Strep-Tactin®XT allows in average the purification of a 2-fold higher protein yield compared to Strep-Tactin®.

Strep-Tactin®XT allows intensive washing without loss in yield

An advantage ot the new Strep-tag® system III is that it allows intensive washing without the loss in protein yield. This figure shows the purification of mCherry-Twin-Strep-tag® fusion protein via 1 ml Strep-Tactin® Superflow® resin and 1 ml Strep-Tactin®XT Superflow® resin. E. coli supernatants were prepared and applied onto the column. Then the columns were washed with 8 CV Buffer W in order to reach a constant absorption at A260/A280 nm. Then the target protein was eluted with 50 mM biotin. Samples of the last wash fraction and the elution fraction were analysed on SDS-PAGE showing that some of the target protein was washed down from Strep-Tactin® Superflow® resin, whereas no protein was eluted from Strep-Tactin®XT Superflow® resin.

The elution fractions were further analysed in Western Blot, showing that the remaining bands are degradation products of mCherry.

Efficient immobilization of Strep-tag® fusion proteins via Strep-Tactin®XT

The following assay shows the comparison of the immobilization efficiency of bacterial alkaline phosphatase (BAP-Strep-tag®II) on Strep-Tactin®XT and Strep-Tactin®. Here, different amounts of BAP were subjected to the coated microplates. After washing, the relative maximum amount of immobilized BAP was determined. The relative signal intensity reflects the amount of bound BAP.

Download complete data set

Get an overview on the system components

Two different affinity tags

The Strep-tag® system is based on one of the strongest non-covalent interactions in nature, which is the interaction of biotin to streptavidin. It allows the purification, detection and immobilization of highly pure and bioactive recombinant proteins. The system includes two affinity tags: Strep-tag®II is a synthetic peptide consisting of eight amino acids and Twin-Strep-tag® consists of two Strep-tag®II sequences in series combined by a GS-linker. This peptide sequences exhibit intrinsic affinity towards Strep-Tactin® and Strep-Tactin®XT, two specifically engineered streptavidin variants.

Two different resins

 

Strep-Tactin® and Strep-Tactin®XT are engineered Streptavidin variants exhibiting different binding affinities towards Strep-tag® and Twin-Strep-tag®.