SPYCATCHER2

100% Secure


SpyCatcher2

SpyCatcher2-CYS

Product Type
Recombinant Protein
Product Code Applications Pack Size List Price Quantity
TZC001
Datasheet
IY
SDS
1 mg loader

TZC001CYS
Datasheet
IY
SDS
1 mg loader

SpyCatcher2 (H-SpyC2) is a 15.7 kDa protein that forms a stable covalent isopeptide bond with a second protein that includes a SpyTag. The reaction occurs spontaneously on mixing, is rapid, irreversible, high-yielding and shows good specificity. SpyCatcher2 contains a His6-tag at the N-terminus. To avoid deamidation of asparagine, a N105D mutation was introduced (Hentrich et al. 2020).

The reaction is robust at pH 5 to 8, at temperatures from +4°C to +37°C, in various buffer conditions (Ca2+/Mg2+ not needed) and in the presence of detergents. The reaction also occurs inside cells (in vivo). Since the speed of the coupling reaction is concentration dependent, it is recommended to use SpyCatcher2 undiluted.

All the different formats of SpyCatcher2 are compatible with Bio-Rad's recombinant HuCAL® antibodies with a SpyTag2 at the C-terminus of the heavy chain.

Download SpyCatcher coupling protocol

SpyCatcher2 (TZC001) is available as a modified version, SpyCatcher2-CYS (TZC001CYS), which contains a cysteine residue for site-specific conjugation by the user.

SpyCatcher2-CYS can dimerize during storage by formation of a disulfide bond via the free cysteines. Before conjugation to these cysteines, the SpyCatcher2-CYS dimers should be mildly reduced e.g., by addition of 5mM DTT and incubation for 1 hr at room temperature, followed by a fast DTT removal step, e.g., by size exclusion chromatography using PD-10 desalting columns, as DTT can interfere with the subsequent conjugation chemistry. Dialysis is not recommended for DTT removal, as the cysteines can oxidize again during this slow process.

View all available Catchers

Product Details

Target Species
Protein/peptide tag
Product Form
SpyCatcher2 purified recombinant protein - liquid
Product Form
SpyCatcher2 purified recombinant protein, with an engineered cysteine residue; use for site-specific chemical conjugation to a label of choice - liquid
Preparation
Recombinant protein expressed in E. coli and purified by affinity chromatography
Preparation
Recombinant protein expressed in E. coli and purified by affinity chromatography
Buffer Solution
Phosphate buffered saline
Buffer Solution
Phosphate buffered saline
Preservative Stabilisers
0.1% ProClin 950
Preservative Stabilisers
0.1% ProClin 950
5 mM EDTA
Purity
≥90% determined by SDS-PAGE under reducing conditions and visualized by coomassie blue staining
Purity
≥90% determined by SDS-PAGE under reducing conditions and visualized by coomassie blue staining
Approx. Protein Concentrations
250 µM (3.9 mg/ml)
Approx. Protein Concentrations
440 µM (7 mg/ml)
Protein Molecular Weight
Predicted 15687 Da

Storage Information

Storage
Store at -20°C only.
Storage in frost-free freezers is not recommended.
This product should be stored undiluted. Avoid repeated freezing and thawing as this may denature the protein. Should this product contain a precipitate we recommend microcentrifugation before use.
Storage
Store at -20°C only.
Storage in frost-free freezers is not recommended.
This product should be stored undiluted. Avoid repeated freezing and thawing as this may denature the protein. Should this product contain a precipitate we recommend microcentrifugation before use.
Guarantee
12 months from date of despatch
Guarantee
12 months from date of despatch

More Information

Acknowledgements
ProClin is a trademark of The Dow Chemical Company (“Dow”) or an affiliated company of Dow
Sold under license of patent and patent applications, including but not limited to the international PCT application PCT/GB2018/051065, the European patent EP2534484B1, UK patent GB1706430.4, the US patents US9,547,003, US10,247,727, US patent application US16/274908 and corresponding patents
HuCAL® is a registered trademark of MorphoSys AG
Acknowledgements
ProClin is a trademark of The Dow Chemical Company (“Dow”) or an affiliated company of Dow
Sold under license of patent and patent applications, including but not limited to the international PCT application PCT/GB2018/051065, the European patent EP2534484B1, UK patent GB1706430.4, the US patents US9,547,003, US10,247,727, US patent application US16/274908 and corresponding patents
HuCAL® is a registered trademark of MorphoSys AG
Licensed Use
For in vitro research purposes. Any re-sale in any form or any other commercial application needs a written agreement with Bio-Rad.
Regulatory
For research purposes only

Applications of SPYCATCHER2

This product has been reported to work in the following applications. This information is derived from testing within our laboratories, peer-reviewed publications or personal communications from the originators. Please refer to references indicated for further information. For general protocol recommendations, please visit the antibody protocols page.
Application Name Verified Min Dilution Max Dilution
Immunoassay
Immunoassay
Where this product has not been tested for use in a particular technique this does not necessarily exclude its use in such procedures. Suggested working dilutions are given as a guide only. It is recommended that the user titrates the product for use in their own system using appropriate negative/positive controls.
Where this product has not been tested for use in a particular technique this does not necessarily exclude its use in such procedures. Suggested working dilutions are given as a guide only. It is recommended that the user titrates the product for use in their own system using appropriate negative/positive controls.
Instructions For Use
View SpyCatcher Coupling Protocol
Instructions For Use
View SpyCatcher Coupling Protocol

Useful Reagents Available

Description Product Code Applications Pack Size List Price Quantity
SpyCatcher2-CYS TZC001CYS IY 1 mg loader
SpyCatcher2 TZC001 IY 1 mg loader

Product Specific References

Antibody Characterization Reference

  1. Hentrich, C. et al. (2020) Periplasmic Expression of SpyTagged Antibody Fragments Enables Rapid Modular Antibody Assembly
    [Epub ahead of print] [Epub ahead of print]
  2. Zakeri, B. et al. (2012) Peptide tag forming a rapid covalent bond to a protein, through engineering a bacterial adhesin.
    Proc Natl Acad Sci U S A. 109 (12): E690-7. Proc Natl Acad Sci U S A. 109 (12): E690-7.
  3. Keeble, A.H. et al. (2017) Evolving Accelerated Amidation by SpyTag/SpyCatcher to Analyze Membrane Dynamics.
    Angew Chem Int Ed Engl. 56 (52): 16521-16525. Angew Chem Int Ed Engl. 56 (52): 16521-16525.

Fluorescent Spectraviewer

Watch the Tool Tutorial Video ▸

How to Use the Spectraviewer

Watch the Tool Tutorial Video ▸
  • Start by selecting the application you are interested in, with the option to select an instrument from the drop down menu or create a customized instrument
  • Select the fluorophores or fluorescent proteins you want to include in your panel to check compatibility
  • Select the lasers and filters you wish to include
  • Select combined or multi-laser view to visualize the spectra