Fluorescence‐labeled peptides as isoelectric point (p I ) markers in capillary isoelectric focusing with fluorescence detection

Commercially available peptides, mostly angiotensin derivatives, were labeled at their N ‐terminal amino group with 5‐carboxytetramethylrhodamine succinimidyl ester, to obtain fluorescent p I markers for capillary isoelectric focusing with fluorescence detection. The labeled peptides were purified by reversedphase chromatography. They were well separated on isoelectric focusing in a polyacrylamide gel slab and their p I s were determined by comigration with protein‐pI markers. The fluorescent markers could be detected as sharp peaks in capillary isoelectric focusing with laser‐induced fluorescence detection (He‐Ne laser, 1 mW, 543.5 nm). The detection limit was found to be around 3 × 10 −12 M (0.8 amol). Tetramethylrhodamine‐labeled pea lectin (3 pg) was subjected to capillary isoelectric focusing and the p I s of the fluorescent derivatives of the lectin were determined by using the fluorescence‐labeled peptides as p I markers.