SNAP-Cell® Oregon Green® is a photostable green fluorescent substrate that can be used to label SNAP-tag® fusion proteins inside living cells or in vitro. Conjugates of Oregon Green are more photostable than those of fluorescein, and their fluorescence properties are essentially pH insensitive in the physiological pH range.
It is a cell-permeable substrate (BG-Oregon Green) based on the Invitrogen dye, Oregon Green
It is suitable for standard fluorescein filter sets
It has an excitation maximum at 490 nm and an emission maximum at 514 nm
Product Information
This package contains 50 nmol of SNAP-Cell Oregon Green substrate, sufficient to make 10 ml of a 5 µM SNAP-tag fusion protein labeling solution.
The SNAP-tag is a novel tool for protein research, allowing the specific, covalent attachment of virtually any molecule to a protein of interest. The SNAP-tag is based on mammalian O6-alkylguanine-DNA-alkyltransferase (AGT). SNAP-tag substrates are derivatives of benzylguanines and benzylchloropyrimidines. In the labeling reaction, the dye-substituted benzyl group of the substrate becomes covalently attached to the SNAP-tag.
There are two steps to using this system: subcloning 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.
Figure 1: Live U-2 OS cells transiently transfected with pSNAP-Cox8A (mitochondrial expression).
Cells were labeled with SNAP-Cell Oregon Green (green) for 15 minutes and counterstained with Hoechst 33342 (blue).
Figure 2:
Excitation (dotted line) and emission spectra after coupling of SNAP-Cell Oregon Green to SNAP-tag in buffer at pH 7.5.
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
S9104S
-20
SNAP-Cell® Oregon Green®
S9104SVIAL
-20
1 x 50 nmol
Not Applicable
Properties & Usage
Materials Required but not Supplied
Cells expressing SNAP-tag fusion proteins
Tissue culture materials and media
Transfection reagents
Fluorescence microscope with suitable filter set
DMSO
Excitation
490nm
Emission
514nm
Product Notes
Storage: SNAP-Cell Oregon Green 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.
SNAP-Cell Oregon Green is not suitable for labeling of cell surface SNAP-tag fusion proteins. The Oregon Green dye is based on a fluorinated fluorescein modified with pivaloyl groups which is uncharged and essentially non-fluorescent. It becomes fluorescent once inside the cell where it is hydrolyzed by non-specific esterases.
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).
After the wash steps, incubate the intracellular labeling samples at 37°C, 5% CO2 for 30 extra minutes to allow any unincorporated fluorophore to diffuse out of the cells.
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
Notice To Buyer/User: The Buyer/User has a non-exclusive license to use this system or any component thereof for RESEARCH AND DEVELOPMENT PURPOSES ONLY. Commercial use of this system or any components thereof requires a license from New England Biolabs, Inc., 240 County Road, Ipswich, MA 01938.
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.
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.
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 This product is provided under an intellectual property license from Life Technologies Corporation. The purchase and/or transfer of this product conveys to the buyer the non-transferable right to use the purchased amount of the product and components of the product only in research for either directed covalent labeling of proteins conducted by the buyer (whether the buyer is an academic or for-profit entity), according to the product instructions. The buyer must not use the product or its components (1) in manufacturing; (2) to provide a service, information, or data; (3) for therapeutic, diagnostic or prophylactic purposes; (4) resale, whether or not such product or its components are resold for use in research; or for any other commercial purpose. For information on purchasing a license to this product for purposes other than research, contact Life Technologies Corporation, 5791 Van Allen Way, Carlsbad, CA 92008, USA or outlicensing@lifetech.com. Oregon Green™ is a trademark owned by Life Technologies Corporation
The products and/or their use may be covered by one or more of the following patents and patent applications: Methods for Using O6-Alkylguanine-DNA-Alkyltransferases: US 7,939,284; US 8,367,361; EP 1 410 023; EP 2 037 271; EP 1 696 234; EP 2 211 177; JP 4195815; JP 4226053; CN 1295510; CN 1975422; and SG 100125.
Substrates for O6-Alkylguanine-DNA Alkyltransferase: US 7,799,524; and JP 4976651.
Mutants of O6-Alkylguanine-DNA-Alkyltransferases: US 7,888,090. EP 1720981 (pending).
Specific Substrates for O6-Alkylguanine-DNA-Alkyltransferases: US 8,163,479. EP 1730298 (pending).
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