CLIP-Surface™ 547 is a photostable red fluorescent substrate that can be used to label CLIP-tag™ fusion proteins on the surface of living cells or in vitro.
This is a cell impermeable substrate (BC-547) based on the Dyomics dye DY-547
It is suitable for use with standard TAMRA or Cy3 filter sets
It has an excitation maximum at 554 nm and emission maximum at 568 nm
Product Information
This package includes 50 nmol of CLIP-Surface 547 substrate, sufficient to make 10 ml of a 5 µM CLIP-tag fusion protein labeling solution.
The CLIP-tag protein labeling system enables the specific, covalent attachment of virtually any molecule to a protein of interest. CLIP-tag is a protein tag based on human O6-alkylguanine-DNA-alkyltransferase (hAGT). CLIP-tag substrates are derivatives of benzylcytosine (BC). In the labeling reaction, the substituted benzyl group of the substrate is covalently attached to the reactive cysteine of CLIP-tag forming a stable thioether bond. Although CLIP-tag is based on the same protein as SNAP-tag®, the benzylcytosine substrates form a separate class of substrates, different from the benzylcytosine substrates form a separate class of substrates, different from the benzylguanine substrates recognized by SNAP-tag. CLIP-tag and SNAP-tag can be used for orthogonal and complementary labeling of two proteins simultaneously in the same cells.
There are two steps to using this system: subcloning and expression of the protein of interest as a CLIP-tag fusion, and labeling of the fusion with the CLIP-tag substrate of choice. Expression of CLIP-tag fusion proteins is described in the documentation supplied with CLIP-tag plasmids. The labeling of the fusion proteins on the cell surface with the CLIP-tag substrate is described in this document.
Figure 1:Live CHO-K1 cells transiently transfected with pCLIP-NK1R Cells were labeled with CLIP-Surface 547 (red) for 60 minutes and counterstained with Hoechst 33342 (blue).
Figure 2: Excitation (dotted line) and emission (full line) spectra of CLIP-Surface 547 coupled to CLIP-tag in buffer at pH 7.5 Figure 3. Structure of CLIP-Surface 547 (MW 851.0 g/mol)
This product is related to the following categories:
The following reagents are supplied with this product:
NEB #
Component Name
Component #
Stored at (°C)
Amount
Concentration
S9233S
-20
CLIP-Surface™ 547
S9233SVIAL
-20
1 x 50 nmol
Not Applicable
Properties & Usage
Materials Required but not Supplied
Cells expressing CLIP-fusion proteins. The protein of interest can be expressed with the CLIP-tag as either an N- or a C-terminal fusion, but note that the tag needs to be exposed to the extracellular surface of the plasma membrane for labeling with CLIP-Surface 547.
Tissue culture materials and media
Transfection reagents
Fluorescence microscope with suitable filter set
DMSO
Excitation
554nm
Emission
568nm
Product Notes
Storage: CLIP-Surface 547 should be stored at -20°C (long term) or at 4°C in the dark (short term, less than 4 weeks). Protect the substrate from light and moisture. With proper storage at -20°C the substrate should be stable for at least three years dry or 3 months dissolved in DMSO.
After addition of DMSO, pipette up and down at least 10-20 times and vortex vigorously for at least one full minute to ensure full dissolution of the substrate.
After diluting the substrate in complete medium, thoroughly pipette up and down at least 10 times to help reduce background.
Increasing substrate concentration and/or reaction time usually results in higher background and does not necessarily increase the signal-to-background ratio (SNR).
Citations & Technical Literature
Citations
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Quality, Safety & Legal
Quality Control Assays
Quality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.
Specifications & Change Notifications
The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]
The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]
Products and content are covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB). The use of trademark symbols does not necessarily indicate that the name is trademarked in the country where it is being read; it indicates where the content was originally developed. The use of this product may require the buyer to obtain additional third-party intellectual property rights for certain applications. For more information, please email busdev@neb.com.
This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.
New England Biolabs (NEB) is committed to practicing ethical science – we believe it is our job as researchers to ask the important questions that when answered will help preserve our quality of life and the world that we live in. However, this research should always be done in safe and ethical manner. Learn more.
Licenses
Cellular Imaging and Analysis (i.e., SNAP and CLIP products)
Notice to Buyer/User: The Buyer/User has a non-exclusive license to use this system or any component thereof for RESEARCH AND DEVELOPMENT ONLY. Commercial use of this system or any components thereof requires a license from New England Biolabs, Inc., 240 County Road, Ipswich, MA 01938. For detailed information see our Terms of Use .
These patents and patent applications are owned by Covalys, or owned by the Ecole Polytechnique Fédérale de Lausanne (EPFL) and exclusively licensed to Covalys and NEB.
The products and/or their use may be covered by one or more of the following patents and patent applications: Labeling of Fusion Proteins with Synthetic Probes: US 8,227,602; US 8,623,627; and EP 2 049 499.
