Streptactin(Strep-Tag II) Resin 4FF

Product Overview

Streptactin (Strep-Tag II) Resin 4FF is a 4% highly cross-linked agarose affinity medium functionalized with Strep-Tactin for selective purification of proteins carrying Strep-tag II (WSHPQFEK) or tandem/Twin Strep-tag sequences. It enables >95% purity in a single step with gentle desthiobiotin/biotin elution under near-physiological conditions, maintains protein activity, tolerates high flow rates, and supports gravity, batch, and FPLC workflows across bacterial, yeast, insect, and mammalian expression systems. For research use only.

Features

• High specificity – strong affinity for Strep-tag II and Twin-Strep-tag® fusion proteins with minimal nonspecific binding.

• Mild elution – proteins are eluted with desthiobiotin or biotin under near-physiological conditions, preserving protein structure and activity.

• High purity in one step – >95–99% purity typically achieved from crude lysates in a single purification step.

• Stable & reusable – 4% highly cross-linked agarose (4FF) provides excellent mechanical stability, enabling high flow rates and multiple regeneration cycles.

• Broad compatibility – resistant to a wide range of buffers, detergents, denaturants, and reducing agents.

• Flexible format – suitable for gravity flow, batch binding, and automated FPLC systems.

Specification

Performance

• Delivers high recovery and purity of recombinant proteins from bacterial, yeast, insect, and mammalian expression systems.

• Suitable for both low-abundance targets and high-molecular-weight proteins, due to large pore size and optimized ligand density.

• Typical results: >95% protein purity in one step with preserved biological activity.

• Supports repeated cycles of protein purification with consistent binding capacity.

Principle

Strep-tag II is a small peptide tag (WSHPQFEK, ~1 kDa) genetically fused to the protein of interest.
The resin is functionalized with Strep-Tactin®, a streptavidin variant engineered for higher affinity and specificity to the Strep-tag.
Fusion proteins bind selectively to the resin, while untagged proteins are washed away.
Elution is performed using desthiobiotin (a biotin analog) or biotin, which competes for the binding sites, releasing the pure protein under gentle, physiological conditions.

Procedure

1. Equilibration – wash resin with binding buffer.

2. Binding – apply clarified lysate containing Strep-tag fusion proteins.

3. Wash – remove unbound or nonspecific proteins with wash buffer.

4. Elution – add buffer containing desthiobiotin or biotin to release target protein.

5. Regeneration – wash with regeneration buffer (e.g., 2–3 column volumes of 0.1 M NaOH or appropriate cleaning buffer).

6. Reuse – resin can be used for at least 50 cycles with proper regeneration.

Reagents Compatible with the Strep-tag/Strep-Tactin Interaction

• Detergents: Triton X-100, Tween 20, NP-40, CHAPS, digitonin, n-dodecyl maltoside

• Reducing agents: DTT, β-mercaptoethanol

• Salts: NaCl (up to 5 M), KCl, CaCl₂, MgCl₂

• Denaturants: Urea, guanidine-HCl (compatible at moderate concentrations)

• Others: glycerol, ethanol, imidazole, PMSF
  ⚠️ High concentrations of free biotin should be avoided as they irreversibly occupy binding sites.

Applications

• Purification of Strep-tag II and Twin-Strep-tag® recombinant proteins from various expression systems (bacteria, yeast, insect, mammalian).

• Isolation of protein complexes and protein–protein interaction studies (pull-down assays).

• Preparation of functional proteins for structural biology (X-ray crystallography, cryo-EM) and biophysical assays.

• Enzyme purification for activity assays and drug discovery.

• Sequential two-tag purification (e.g., His-tag + Strep-tag) for ultrapure proteins.

• Suitable for both research-scale and preparative-scale protein production.

Q&A for Streptactin (Strep-Tag II) Resin 4FF

Q1: What types of proteins can be purified with Streptactin (Strep-Tag II) Resin 4FF?
A1: The resin is designed for recombinant proteins carrying a Strep-tag II or Twin Strep-tag. It works efficiently with proteins expressed in bacteria, yeast, insect, or mammalian cells, and is suitable for both small and high-molecular-weight targets.

Q2: How is the protein eluted from the resin?
A2: Proteins are typically eluted under mild conditions using desthiobiotin or biotin. These competitive ligands preserve protein activity and structure, making the resin ideal for sensitive proteins.

Q3: What is the binding capacity of this resin?
A3: The typical binding capacity is about 4 mg of Twin Strep-tag fusion protein per mL of resin. The exact capacity depends on protein size, expression system, and buffer conditions.

Q4: Can the resin be reused?
A4: Yes. With proper regeneration (e.g., mild NaOH wash), the resin can be reused for at least 50 purification cycles while maintaining consistent performance.

Q5: Is the resin compatible with detergents and additives?
A5: Yes. The Strep-tag/Streptactin interaction is stable in the presence of detergents (Triton X-100, Tween 20, CHAPS, etc.), salts, reducing agents (DTT, β-mercaptoethanol), and even moderate concentrations of denaturants (urea, guanidine-HCl).

Q6: How does Strep-tag purification compare with His-tag purification?
A6: Strep-tag purification typically yields higher purity in a single step, with milder elution conditions that help preserve protein function. Unlike IMAC (His-tag/Ni-NTA), it is not affected by EDTA or metal chelators in the buffer.

Q7: What storage conditions are recommended for the resin?
A7: Store at 2–8 °C in PBS containing 20% ethanol. Do not freeze. Under these conditions, the resin is stable for long-term use.

Storage

2~8°C, do not freeze