The SNAP-tag®, CLIP-tag™, and ACP-tag/MCP-tag protein labeling technologies offer an innovative alternative to traditional fluorescent proteins for studying the function and localization of proteins in living and fixed cells. Covalent protein labeling brings simplicity and versatility to the imaging of mammalian proteins in live cells, as well as the ability to capture proteins in vitro. The creation of a single genetic construct generates a fusion protein which, when covalently attached to a variety of substrates, including fluorophores, biotin, and beads, provides a powerful tool for studying proteins. In the SNAP-tag and CLIP-tag sytems, the fusion protein is labeld by a self-labeling tag based on the DNA repair protein O6-alkylguanine-DNA-alkyltrasferase, whereas with the ACP-tag and MCP-tag systems, the labeling is catalyzed through a post-translational modification by a phosphopantetheinyl transferase.
Another way of labeling proteins is by using intein-mediated protein ligation (1) or expressed protein ligation. The IPL reaction allows the ligation of a synthetic peptide or a protein with an N-terminal cysteine residue to the thioester on the C-terminus of an expressed protein through a native peptide bond. The IPL protocol employs an IMPACT™ (NEB #E6901) C-terminal fusion vector to express and purify a protein of interest and to generate a thioester at its C-terminus. IPL allows for ligation of peptides (may contain modified amino acids) or proteins to the C-terminus of an IMPACT expressed protein.
- 5 Minute Transformation Protocol (C3022)
- Affinity Purification and On-column Cleavage (E6901)
- Cellular Labeling (E9100)
- Cellular Labeling (E9230)
- Cellular Labeling (S9124)
- Cloning of ACP-tag Fusions in pACP-tag(m)-2 (N9322)
- Construction of the Fusion Plasmid (E6901)
- Expression of ACP-tag Fusions (N9322)
- Fusion Protein Expression (E6901)
- High Efficiency Transformation Protocol (C3022)
- Labeling of Proteins in Solution (E9230)
- Labeling of Proteins in vitro (S9351)
- Labeling on the Surface of Cells (S9349)
- Labeling on the Surface of Cells (S9350)
- Labeling on the Surface of Cells (S9351)
- Labeling Proteins in vitro (S9124)
- Labeling SNAP-tag Purified Protein In Vitro (P9312)
- Preparation of Media and Solutions (E6901)
- Primer Design for Restriction Enzyme Cloning (E6901)
- Protocol for Expression Using T7 Express Crystal (C3022)
- Recommended media and expression conditions for T7 Express Crystal (C3022)
- Seleno-methionine Incorporation (C3022)
- Simplified Expression and Purification Protocol (E6901)
- Use of SNAP-Cell Block with SNAP-Cell Substrates (E9100)
- Use of CLIP-Cell Block with CLIP-Cell Substrates (E9230)
- View the video "Fluorescent Labeling of COS-7 Expressing SNAP-tag Fusion Proteins for Live Cell Imaging" in the Journal of Visualized Experiments (JoVE)
- Western Transfer Protocol for Anti-MBP Monoclonal Antibody
- Cellular Labeling (E9120)
- Instructions for Cellular Labeling (E9200)
- Labeling Proteins in vitro (E9120)
- Labeling Proteins in vitro (E9200)
- Protocol for IPL
- Reaction Conditions for Chemical Coupling with CoA-SH (S9352S)
- Cell Surface Labeling ACP-Surface Starter Kit
- Labeling of Proteins in vitro for ACP-Surface Starter Kit
- Cloning of SNAP-tag Fusions in pSNAPf (N9183)
- Cloning of SNAP-tag Fusions in pSNAP-tag(T7)-2 (N9181)
- Expression of SNAP-tag Fusions (N9181)
- Expression of SNAPf Fusions (N9183)
- Labeling of Proteins in vitro (S9349)
- Labeling of Proteins in vitro (S9350)
- Protocol for Labeling ACP- or MCP-tag Fusion Proteins with CoA Substrates.
- Labeling of Proteins in vitro (E9100)
- Use SNAP-Capture Magnetic Beads (S9145)
- RNase B Deglycosylation Protocol (P7817)
- Fusion Constructs (E6901)
- Intein-Mediated Protein Ligation (IPL) and Labeling with the IMPACT™ Kit
- Labeling and Imaging of Cell Surface Receptors Mediated by SNAP-tag®
- Labeling of Escherichia coli Expressed SNAP-tag Fusion Proteins C2566
- Simultaneous Labeling of Two Proteins in Live Cells S9141
- SNAPf based pulse labeling for analysis of protein turnover in living cells
SNAP-tag® Technologies: Tools to Study Protein Function
Read about the NEB’s set of protein tools for the specific labeling (SNAP-, CLIP-, ACP- and MCP-tags) of fusion proteins.
- Cellular Imaging & Analysis Brochure
- Comparison of SNAP-tag®/CLIP-tag™ Technologies to GFP
- Labeling with SNAP-tag® Technology Troubleshooting Guide
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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.