Thermo Scientific™ Pierce™ Cell Surface Protein Isolation Kit

Catalog No.	Quantity
PI89881	8 reactions kit

Catalog No. PI89881 Supplier Thermo Scientific™ Supplier No. 89881

Description

Selectively biotinylate and purify mammalian cell surface proteins and exclude intracellular proteins with this complete set of reagents.

Thermo Scientific™ Cell Surface Protein Isolation Kit is a complete set of reagents for selective biotinylation and subsequent purification of mammalian cell surface proteins to the exclusion of intracellular proteins.

The kit efficiently labels proteins that have accessible lysine residues and sufficient extracellular exposure.The isolation procedure uses a cell-impermeable, cleavable biotinylation reagent (Sulfo-NHS-SS-Biotin) to label exposed primary amines of proteins on the surface of intact adherent or suspension cells. Treated cells are then harvested and lysed, and the labeled surface proteins are affinity-purified using Thermo Scientific NeutrAvidin Agarose Resin. The isolated cell surface proteins contain a small, nonreactive tag of the originally labeled primary amines but are no longer biotinylated (biotin remains bound to the resin). The kit contains sufficient reagents for eight experiments, each involving four T75 flasks of confluent cells.

Highlights:

• Isolates cell surface proteins – reduces complexity of total cellular protein

• Efficiently recovers labeled proteins – cleavable biotin allows for nearly 100% recovery of isolated cell surface proteins

• Convenience – all reagents are provided in one kit, along with complete instructions for labeling, cell lysis and purification of cell surface membrane proteins

• Western blotting applications – proteins recovered in SDS-PAGE buffer are loaded directly onto polyacrylamide gels

• Robust system – protocol designed for diverse cell lines, including NIH 3T3, HeLa, C6 and A431

Includes:

Biotinylation reagent, cell lysis reagent, agarose affinity resin, various buffers and microcentrifuge spin columns

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Specifications

• Product Type: Protein Isolation Kit
• Content And Storage: Upon receipt store kit at 4°C. Product shipped at ambient temperature.
• Form: Liquid, Powder
• Product Line: Pierce
• Buffer: Lysis Buffer
• Reagent Type: Cell Lysis Buffer, Detergent Solution, Subcellular Fractionation
• Sufficient For: 8 Reactions
• Quantity: 8 reactions kit

Frequently Asked Questions (FAQs)

Can you provide the shelf-life for the Pierce Cell Surface Protein Isolation Kit?

The Pierce Cell Surface Protein Isolation Kit is covered under our general 1-year warranty and is guaranteed to be fully functional for 12 months from the date of shipment, if stored as recommended. Please see section 8.1 of our Terms & Conditions of Sale (https://www.thermofisher.com/content/dam/LifeTech/Documents/PDFs/Terms-and-Conditions-of-Sale.pdf) for more details.

Can I use conjugated streptavidin to detect proteins isolated with the Thermo Scientific Cell Surface Protein Isolation Kit following my Western blot transfer?

No. As a cleavable biotin is used to release the labeled proteins from the NeutrAvidin Agarose in a relatively gentle manner, incubation of the labeled proteins on the NeutrAvidin Agarose with a reducing agent will cleave the disulfide bond in the spacer arm of the biotinylation reagent, leaving the biotin on the NeutrAvidin Agarose, and allowing the protein to be recovered free of the biotin tag.

If I want to obtain a higher level of labeling of my cell surface molecules is it okay to increase the concentration of the Sulfo-NHS-SS-Biotin?

No. Biotin is a small compound and high concentrations can behave similar to high concentrations of salt, compromising the cell membrane and allowing proteins to escape from the cell's interior. We do not recommend using a concentration higher than 0.25 mg/mL of the Sulfo-NHS-SS-Biotin when labeling surface proteins.

Why does the Thermo Scientific Cell Surface Protein Isolation Kit include a cleavable version of water soluble biotin?

The biotinylation reagent is water soluble to prevent it from passing through the cell membrane and labeling proteins on the interior of the cell. This reagent is cleavable in order to allow release from the NeutrAvidin Agarose. NeutrAvidin, which is deglycosylated avidin, has the same binding affinity for biotin as does avidin (Kd = 1.3 x 10-15 at pH 5), which is often considered the strongest non-covalent interaction known. Conditions required to release the biotin from the NeutrAvidin do so by irreversibly denaturing the NeutrAvidin protein, as well as most other proteins. Cleavage via reduction with a sulfhydryl-containing compound is a much gentler recovery method for the labeled cell surface proteins.

Safety and Handling

WARNING: Cancer - www.P65Warnings.ca.gov

For Research Use Only. Not for use in diagnostic procedures.