Labeling Small Peptides with Amine-Reactive Dyes in Organic Solvents—Note 9.2
Most of the product literature associated with our amine-reactive dyes provides protocols for labeling proteins, typically IgG antibodies in aqueous buffers (Amine-Reactive Probe Labeling Protocol). The following protocol is a starting point for labeling peptides in organic solvents. Please note that the reaction conditions, including concentrations of the reactants and the reaction times, may require optimization. Furthermore, many peptides are not soluble in a 100% organic solution. It is very important to test the solubility of the peptide in dimethylsulfoxide (DMSO) or dimethylformamide (DMF) before attempting this procedure.
- Dissolve the peptide to be labeled in DMSO or DMF at 0.1–1 mM.
- Add 100 mM triethylamine to the reaction solution. This will ensure that the amines to be derivatized are deprotonated.
- Add the amine-reactive dye to the reaction solution. The reactive dye should be in a 1:1 to 3:1 molar ratio to the peptide.
- React at room temperature or at 4°C for at least 4 hours with continuous stirring, protected from light. The reaction can proceed overnight. Thin-layer chromatography may be useful for monitoring the reaction's progress.
- Purify the conjugate by an appropriate method, such as HPLC-based separation.