SNAP- and CLIP-tag protein labeling systems enable the specific, covalent attachment of virtually any molecule to a protein of interest. There are two steps to using this system: cloning and expression of the protein of interest as a SNAP-tag® fusion, and labeling of the fusion with the SNAP-tag substrate of choice. The SNAP-tag is a small protein based on human O6-alkylguanine-DNA-alkyltransferase (hAGT), a DNA repair protein. SNAP-tag substrates are dyes, fluorophores, biotin, or beads conjugated to guanine or chloropyrimidine leaving groups via a benzyl linker. In the labeling reaction, the substituted benzyl group of the substrate is covalently attached to the SNAP-tag. CLIP-tag™ is a modified version of SNAP-tag, engineered to react with benzylcytosine rather than benzylguanine derivatives. When used in conjunction with SNAP-tag, CLIP-tag enables the orthogonal and complementary labeling of two proteins simultaneously in the same cells.
SNAP-tag® is a registered trademark of New England Biolabs, Inc.
CLIP-tag™ is a trademark of New England Biolabs, Inc.
Cellular Analysis includes these subcategories:
FAQs for Cellular Analysis
SNAP-tag® Technologies: Novel Tools to Study Protein Function
Cellular Imaging & Analysis Brochure
The Cellular Imaging and Analysis brochure provides information on the labeling technologies offered by NEB for studying the function and localization of proteins in cells.
- Comparison of SNAP-tag®/CLIP-tag™ Technologies to GFP
- Labeling with SNAP-tag® Technology Troubleshooting Guide
Other Tools & Resources
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- Johnsson K. 2009. Visualizing biochemical activities in living cells Nat Chem Biol . 5 , PubMedID: 19148167, DOI:
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- Kapmeier F. et al. 2009. Site-Specific, covalent labeling of recombinant antibody fragments via fusion to an engineered version of 6-O-alkylguanine DNA alkyltransferase Bioconjug Chem. . 23-Apr , PubMedID: 19388673, DOI:
- Stein V. and Hollfeder F. 2009. An efficient method to assemble linear DNA templates for in vitro screening and selection systems Nuc. Acids Res . 37, PubMedID: 19617373, DOI:
- Donovan C. et al. 2009. Characterization and subcellular localization of bacterial flotillin homologue Microbiology . 155 , PubMedID: 19383680, DOI:
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- Carroll C.W. et al. 2009. Centromere assembly requires the direct recognition of CENP-A nucleosomes by CENP-N Nat. Cell Biol. . 11 , PubMedID: 19543270, DOI:
- Foltz D.R. et al. 2009. Centromere-specific assembly of CENP-a nucleosomes is mediated by HJURP Cell . 137 , PubMedID: 19410544, DOI:
- Ahier A. et al. 2009. A new family of receptor tyrosine kinases with a venus flytrap binding domain in insects and other invertebrates activated by aminoacids PLoS One . 4, PubMedID: 19461966, DOI:
- Cornish, V. W. 2009. Fluorescence in living systems: applications in chemical biology Wiley Encyc. of Chem. Biol. . 2 , PubMedID: , DOI:
- Chattopadhaya S. et al. 2009. Expanding the chemical Biologist's tool kit: chemical labelling strategies and its applications Curr. Med. Chem. . 16 , PubMedID: 19903152, DOI:
- Degorce F. et al. 2009. HTRF: A technology tailored for drug discovery - a review of theoretical aspects and recent applications Curr. Chem. Genomics . 3 , PubMedID: 20161833, DOI:
- Samoshkin A. et al. 2009. Human condensin function is essential for centromeric chromatin assembly and proper sister kinetochore orientation PLoS One . 4 , PubMedID: 19714251, DOI:
- Keppler A. et al. 2009. Chromophore-assisted laser inactivation of α- and γ-tubulin SNAP-tag fusion proteins inside living cells ACS Chem. Biol. . 4 , PubMedID: 19191588, DOI:
- Hill Z. B. 2009. A chemical genetic method for generating bivalent inhibitors of protein kinases J. Am. Chem. Soc. . 131, PubMedID: 19391594, DOI:
- Damoiseaux, R. et al 2002. Towards the generation of artificial O6-alkylguanine-DNA alkyltransferases: in vitro selection of antibodies with reactive cysteine residues ChemBioChem . 3, PubMedID: 12325014, DOI:
- Keppler A. et al. 2003. A general method for the covalent labeling of fusion proteins with small molecules in vivo Nature Biotechnology . 21, PubMedID: , DOI:
- Gendreizig, S. et al. 2003. Induced protein dimerizaton in vivo through covalent labeling JACS . 125, PubMedID: 14653715, DOI:
- Gendreizig S. et al. 2003. Covalent labeling of fusion proteins with chemical probes in living cells Chimia . 57, PubMedID: , DOI:
- Juillerat A. et al. 2003. Directed evolution of O6-alkylguanine-DNA alkyltransferase for efficient labeling of fusion proteins with small molecules in vivo Chem. Biol. . 10, PubMedID: , DOI:
- Keppler A. et al. 2004. Labeling of fusion proteins with synthetic fluorophores in live cells PNAS . 101, PubMedID: , DOI:
- Keppler A. et al. 2004. Labeling of fusion proteins of O6-alkylguanine-DNA alkyltransferase with small molecules in vitro and in vivo Methods . 32, PubMedID: 15003606, DOI:
- Kindermann M. et al. 2004. Synthesis and characterization of bifunctional probes for the specific labeling of fusion proteins Bioorg. Med. Chem. Lett. . 14, PubMedID: , DOI:
- Hoskins, A. et al. 2011. Ordered and dynamic assembly of single spliceoseoms Science . 331 , PubMedID: 21393538, DOI:
- Eckhardt, M. et al. 2011. A SNAP-tagged detivative of HIV-1 - A versatile tool to study virus-cell interactions PLoS One . , PubMedID: 21799764, DOI: 10.137/journal. P One .0022007
- Gronemeyer T. et al. 2006. Directed evolution of O6-alkylguanine-DNA alkyltransferase for applications in protein labeling Prot. Eng. Des. Sel. . 19, PubMedID: 12725859, DOI:
- Keppler A. et al. 2006. Fluorophores for live cell imaging of AGT fusion proteins across the visible spectrum BioTechniques . 41, PubMedID: 16925018, DOI:
- Krayl M. et al. 2006. Fluorescence-mediated analysis of mitochondrial preprotein import in vitro Anal. Biochem. . 335, PubMedID: 16750157, DOI:
- Tirat A. et al. 2006. Evaluation of two novel tag-based labeling technologies for site-specific modification of proteins Int. J. Biol. Macromol.. 39, PubMedID: 16503347, DOI:
- Heinis C. et al. 2006. Evolving the substrate specificity of O6 alkylguanine DNA alkyltransferase through loop insertion for applications in molecular imaging ACS Chem Biol. . 1, PubMedID: 17168553, DOI:
- Gronemeyer T. et al. 2006. Adding value to fusion proteins through covalent labeling Curr. Opin. Biotechn. . 16 , PubMedID: 15967656, DOI:
- Tugulu S. et al. 2005. Protein-functionalized polymer brushes Biomacromolecules . 6, PubMedID: 15877383, DOI:
- Cravatt B.F. 2005. Live chemical reports from the cell surface Chem. Biol. . 12, PubMedID: 16183017, DOI:
- Vivero-Pol L. et al. 2005. Multicolor imaging of cell surface proteins J. Am. Chem. Soc. . 127, PubMedID: 16159249, DOI:
- Yin J. et al. 2005. Single-cell FRET imaging of transferrin receptor trafficking dynamics by Sfp-catalyzed, site-specific protein labeling Chem. Biol . 12, PubMedID: 16183024, DOI:
- Yin J. et al. 2005. Labeling proteins with small molecules by site-specific posttranslational modification J Am Chem Soc. 126 , PubMedID: 15212504, DOI:
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- Mosiewicz, K. A. et al. 2010. Phosphopantetheinyl Transferase-Catalyzed Formation of Bioactive Hydrogels for Tissue Engineering J. Am. Chem. Soc. . 132, PubMedID: 20373804, DOI:
- Engin S. et al. 2010. Benzylguanine Thiol self-assembled monolayers for the immobilization of SNAP-tag proteins on microcontact-printed surface structures Langmuir . ASAP, PubMedID: 20369837, DOI:
- Waichman S. et al. 2010. Functional Immobilization and Patterning of Proteins by an Enzymatic Transfer Reaction Anal. Chem. . 82 , PubMedID: 20092261, DOI:
- Zelman-Femiak, M. et al. 2010. Covalent quantum dot receptor linkage via the acyl carrier protein for single-molecule tracking, internalization, and trafficking studies BioTechniques . 49, PubMedID: 20701592, DOI:
- Liu E and Bruner S. D. 2007. Rational manipulation of carrier-domain geometry in nonribosomal peptide synthetases ChemBioChem. . 8, PubMedID: 17335097, DOI:
- Zhou Z. et al. 2007. Genetically encoded short peptide tags for orthogonal protein labeling by Sfp and AcpS phosphopantetheinyl transferases ACS Chemical Biology . 2, PubMedID: 17465518, DOI:
- Generosi J. et al. 2008. Photobleaching-free infrared near-field microscopy localizes molecules in neurons J. App. Phys. . 104, PubMedID: , DOI:
- Schulz C. and Köhn M. 2008. Simultaneous protein tagging in two colors Chemistry & Biology . 15, PubMedID: 18291310, DOI:
- Iversen L. et al. 2008. Templated protein assembly on micro-contact-printed surface patterns. Use of the SNAP-tag protein functionality Langumuir. May 17, PubMedID: 18484753, DOI:
- Kropf M. et al. 2008. Subunit-specific surface mobility of differentially labeled AMPA receptor subunits Eur. J. Cell Biol. . 87, PubMedID: 18547676, DOI:
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- Gautier A. et al. 2009. Selective cross-linking of interacting proteins using self-labeling tags J. Am. Chem. Soc. . 131, PubMedID: 19916541, DOI:
- Gralle M. et al. 2009. Neuroprotective secreted amyloid precursor protein acts by disrupting amyloid precursor protein dimers J. Biol. Chem. . 284, PubMedID: 19336403, DOI:
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- Kindermann M. et al. 2003. Covalent and selective immobilization of fusion proteins JACS . 125, PubMedID: 12822993, DOI:
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Publications related to Cellular Analysis
- Simultaneous dual protein labeling inside live cells
- Protein localization and translocation
- Pulse-chase experiments
- Receptor internalization studies
- Selective cell surface labeling
- Protein pull-down assays
- Protein detection in SDS-PAGE
- Flow cytometry
- High throughput binding assays in microtiter plates
- Biosensor interaction experiments
- FRET-based binding assays
- Single molecule labeling
- Super-resolution microscopy
Selected Publications by Application
Lukinavičius, G. et al. (2015) "Fluorescent labeling of SNAP-tagged proteins in cells" Methods Mol. Biol. 1266, 107-118.
Corrêa Jr., I. R. (2015) "Considerations and protocols for the synthesis of custom protein labeling probes" Methods Mol. Biol. 1266, 55-79.
Corrêa Jr., I. R. (2014) "Live-cell reporters for fluorescence imaging" Curr. Opin. Chem. Biol. 20, 36-45.
Bosch, P. J. et al. (2014) "Evaluation of fluorophores to label SNAP-tag fused proteins for multicolor single-molecule tracking microscopy in live cells" Biophys. J. 107, 803-814.
Smith, B. A. et al. (2013) "Three-color single molecule imaging shows WASP detachment from Arp2/3 complex triggers actin filament branch formation" eLife 2, e01008.
Jaiswal, R. et al. (2013) "The Formin Daam1 and Fascin Directly Collaborate to Promote Filopodia Formation" Curr. Biol. 23, 1373-1379.
Breitsprecher, D. et al. (2012) "Rocket Launcher Mechanism of Collaborative Actin Assembly Defined by Single-Molecule Imaging" Science 336, 1164-1168.
Hoskins, A. A. et al. (2011) "Ordered and dynamic assembly of single spliceosomes." Science 331 (6022), 1289-1295.
Zhao, Z. W. et al. (2014) "Spatial organization of RNA polymerase II inside a mammalian cell nucleus revealed by reflected light-sheet superresolution microscopy" Proc. Natl. Acad. Sci. USA 111, 681-686.
Lukinavičius, G. et al. (2013) "A near-infrared fluorophore for live-cell super-resolution microscopy of cellular proteins" Nat. Chem. 5, 132-139.
Jones, S. A. et al. (2011) "Fast, three-dimensional super-resolution imaging of live cells." Nat. Methods 8, 499-505.
Klein, T. et al. (2011) "Live-cell dSTORM with SNAP-tag fusion proteins." Nat. Methods 8, 7-9.
Pellett, P. A. et al. (2011) "Two-color STED microscopy in living cells." Biomed. Opt. Expr. 2, 2364-2371
Hein, B. et al. (2010) "Stimulated Emission Depletion Nanoscopy of Living Cells Using SNAP-Tag Fusion Proteins." Biophys. J. 98, 158-163.
Tissue and Animal Imaging:
Yang, G. et al. (2015) "Genetic targeting of chemical indicators in vivo" Nat. Methods 12, 137-139.
Kohl, J. et al. (2014) "Ultrafast tissue staining with chemical tags" Proc. Natl. Acad. Sci. USA 111, E3805-E3814.
Ivanova, A. et al. (2013) "Age-dependent labeling and imaging of insulin secretory granules" Diabetes 62, 3687-3696.
Gong, H. et al. (2012) "Near-Infrared Fluorescence Imaging of Mammalian Cells and Xenograft Tumors with SNAP-Tag" PLoS ONE 7(3): e34003.
Bojkowska K. et al. (2011) "Measuring in vivo protein half-life." Chem. Biol. 18, 805-815.
Cell-Surface Protein Labeling and Internalization Analysis:
Bitsikas, V. et al. (2014) "Clathrin-independent pathways do not contribute significantly to endocytic flux" eLife 3, e03970.
Jaensch, N. et al. (2014) "Stable Cell Surface Expression of GPI-Anchored Proteins, but not Intracellular Transport, Depends on their Fatty Acid Structure" Traffic 15, 1305-1329.
Cole, N. B. and Donaldson, J. G. (2012) "Releasable SNAP-tag Probes for Studying Endocytosis and Recycling" ACS Chem. Biol. 7, 464-469.
Rošić, S. et al. (2014) "Repetitive centromeric satellite RNA is essential for kinetochore formation and cell division" J. Cell Biol. 207, 335-349.
Stoops, E. H. et al. (2014) "SNAP-Tag to Monitor Trafficking of Membrane Proteins in Polarized Epithelial Cells" Methods Mol. Biol. 1174, 171-182.
Bordor, D. L. et al. (2012) "Analysis of Protein Turnover by Quantitative SNAP-Based Pulse-Chase Imaging" Curr. Protoc. Cell Biol. 55, 8.8.1-8.8.34.
Register, A. C. et al. (2014) "SH2-Catalytic Domain Linker Heterogeneity Influences Allosteric Coupling across the SFK Family" Biochemistry 53, 6910-6923.
Shi, G. et al. (2012) "SNAP-tag based proteomics approach for the study of the retrograde route" Traffic 13, 914-925.
Bieling, P. et al. (2010) "A minimal midzone protein module controls formation and length of antiparallel microtubule overlaps" Cell 142, 420-432.
Protein-Protein and Protein-Ligand Interactions:
Griss, R. et al. (2014) "Bioluminescent sensor proteins for point-of-care therapeutic drug monitoring" Nat. Chem. Biol. 10, 598-603.
Chidley, C. et al. (2011) "A yeast-based screen reveals that sulfasalazine inhibits tetrahydrobiopterin biosynthesis." Nat. Chem. Biol. 7, 375-383.
Gautier A. et al. (2009) "Selective Cross-Linking of Interacting Proteins using Self-Labeling Tags" J. Am. Chem. Soc. 131, 17954-17962.
Maurel D. et al. (2008) "Cell-surface protein-protein interaction analysis with time-resolved FRET and SNAP-tag technologies: application to GPCR oligomerization." Nat. Methods 5, 561-567.
Protein Labeling with SNAP-tag and CLIP-tag
SNAP-tag®, CLIP-tag™ and ACP/MCP-tag Substrate Selection Chart
Watch as Chris Provost, of New England Biolabs, performs fluorescent imaging of live COS-7 cells expressing SNAP-tag® fusion proteins.
View an interactive tutorial explaining the mechanism of our SNAP-tag® technologies and reagents available for researchers wishing to study the function and localization of proteins in live or fixed cells.