Use of DTT to Cleave Disulfide-Containing Crosslinking Agents

Dissolve a crosslinked protein or peptide that has been conjugated with the use of a disulfide-containing crosslinker at a concentration of l-10mg/ml in 0.01 M sodium phosphate, 0.15M NaCl, pH 7.4. Alternative buffer conditions and pH values may be used, however a pH between 7.0 and 8.1 usually works best. 

The reduction of disulfides by 2-mercaptoethanol proceeds through a mixed disulfide intermediate. 

Protocol for Preparation and Use of a Gram-Negative Bacteria Lysis Buffer 

Prepare a solution consisting of 2.5 ml glycerol, 100 pi of 10 percent Triton X-100 (Thermo Fisher Surfact-Amps X-100), and 10 pi 2-mercaptoethanol. 

Add lOg of wet packed cells to the lysis buffer and stir vigorously for 30 minutes. 

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