INDIRECT FLUOROMETRIC DETECTION OF TRYPTIC DIGESTS SEPARATED BY CAPILLARY ZONE ELECTROPHORESIS

This work demonstrates the analytical utility of indirect fluorescence detection with capillary zone electrophoresis (CZE) for the analysis of trace quantities of macromolecular mixtures. Detection is based upon charge displacement and is not based upon any absorption or emission property of the analyte. No derivatization step is required. Indirect fluorescence is therefore a general detector for electrophoresis. Subfemtomolar quantities of tryptic digest mixtures are separated within three minutes, and reproducible peaks are obtained from the mixtures. Mass limits of detection are 3000 times lower than those of commercial high-performance liquid chromatography (HPLC) UV absorbance detectors and 180 times lower than those of UV absorbance detectors in CZE systems. This separation and detection system should be well suited to analysis of trace quantities of mixtures of peptides.; This work demonstrates the analytical utility of indirect fluorescence detection with capillary zone electrophoresis (CZE) for the analysis of trace quantities of macromolecular mixtures. Detection is based upon charge displacement and is not based upon any absorption or emission property of the analyte. No derivatization step is required. Indirect fluorescence is therefore a general detector for electrophoresis. Subfemtomolar quantities of tryptic digest mixtures are separated within three minutes, and reproducible peaks are obtained from the mixtures. Mass limits of detection are 3000 times lower than those of commercial high-performance liquid chromatography (HPLC) UV absorbance detectors and 180 times lower than those of UV absorbance detectors in CZE systems. This separation and detection system should be well suited to analysis of trace quantities of mixtures of peptides.