Thermo Scientific Bond-Breaker TCEP Solution, Neutral pH is a stable, 0.5M solution of the thiol-free, phosphine-based TCEP compound, useful as a 10X stock for addition to SDS-PAGE sample loading buffers to reduce protein disulfide bonds.
Bond-Breaker TCEP Solution is a potent, odorless, thiol-free reducing agent with broad application to protein and other research involving reduction of disulfide bonds. This product is an effective and convenient replacement for B-mercaptoethanol or DTT in SDS-PAGE sample buffers. The neutral pH of this reagent provides sharp bands and avoids exposing proteins to the strong acid of TCEP•HCl, which can result in hydrolysis and carbohydrate modification.
- Odorless – unlike DTT and β-ME, TCEP is odor-free, contributing to a healthier lab environment
- Efficient – 5 to 50mM TCEP thoroughly reduces most peptide or protein disulfide bonds within a few minutes (i.e., just as effective as DTT)
- Specific – selective and complete reduction of even the most stable water-soluble alkyl disulfides
- Fast – reduces protein disulfides at room temperature and pH 5 in less than five minutes
- Stable – resistant to air oxidation; nonvolatile and nonreactive toward other functional groups found in proteins
- Versatile – reduces peptides and proteins over a broad range of pH, salt, detergent and temperature conditions
- Compatible – removal of the reducing agent is not necessary before most applications, (e.g. histidine-tagged protein purification, maleimide conjugations), because TCEP does not contain sulfhydryl groups
|Protocol for using Thermo Scientific Bond-Breaker TCEP Solution to reduce proteins for SDS-PAGE analysis.
Properties of Thermo Scientific Bond-Breaker TCEP Solution. See TCEP-HCl (Part No. 20490) for chemical properties of tris(2-carboxyethyl)phosphine.
||6.6 ± 0.1
Considerations for use of Bond-Breaker TCEP Solution:
- Reduction occurs over a wide range of pH (pH 4.0-9.0) and temperature (5 to 95°C).
- Most proteins are reduced efficiently without a denaturant. However, adding a denaturant such as guanidine•HCl may aid in exposing internal disulfides to the Immobilized TCEP.
- Urea is not recommended as a denaturant as it forms cyanates that react with sulfhydryl groups.
- Do not allow metals to contact the TCEP solution as this will decrease TCEP activity.
- Including 5 to 20mM EDTA in the sample buffer during reduction helps prevent reoxidation of the sulfhydryl groups by divalent metals such as Zn 2+, Cu 2+ and Mg 2+.
- The reduced sample should be used immediately after reduction because disulfides will reform over time.
- Irsch, T. and Krauth-Siegel, R.L. (2004) Glyoxalase II of African trypanosomes is trypanothione-dependent. J. Biol. Chem.279, 22209-17.
- Schmidt, H. and Krauth-Siegel, R.L. (2003) Functional and physicochemical characterization of the thioredoxin system in Trypanosoma brucei. J. Biol. Chem.278, 46329-36.
Overview of crosslinking and protein modification
Overview of protein electrophoresis
Immobilized TCEP Disulfide Reducing Gel
All Disulfide Reducing Agents
Electrophoresis Sample Loading Buffers