SNAP-Surface® Alexa Fluor® 647 is a photostable fluorescent substrate that can be used to label SNAP-tag® fusion proteins in solution or on the surface of living cells. This cell impermeable substrate is based on Alexa Fluor® 647 and is suitable for standard 635 nm or 650 nm diode laser excitation. It has an excitation maximum at 652 nm and emission maximum at 670 nm. This package includes 50 nmol of SNAP-Surface Alexa Fluor® 647 substrate, sufficient to make 10 ml of a 5 µM SNAP-tag fusion protein labeling solution.
The SNAP-tag protein labeling system enables the specific, covalent attachment of virtually any molecule to a protein of interest. The SNAP-tag is a protein based on human O6-alkylguanine-DNA alkyltransferase (hAGT). SNAP-tag substrates are 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.
There are two steps to using this system: sub-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. Expression of SNAP-tag fusion proteins is described in the documentation supplied with SNAP-tag plasmids. The labeling of the fusion proteins with the SNAP-tag substrate is described below.
Materials Required but not Supplied
•Cells expressing SNAP-Fusion proteins. Proteins of interest can be expressed with the SNAP-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 SNAP-Surface Alexa Fluor® 647.
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