MINIATURIZED PROTEIN MICROSEQUENCER WITH PTH AMINO-ACID IDENTIFICATION BY CAPILLARY ELECTROPHORESIS .1. AN ARGON PRESSURIZED DELIVERY SYSTEM FOR ADSORPTIVE AND COVALENT SEQUENCING

The design of a simple, highly miniaturized instrument for manual micr osequence analysis of proteins and peptides is described. The reaction chamber is made of fused silica capillary tubing with all reagents an d solvents necessary for coupling and cleavage delivered via two valve s and a syringe-based dispenser. Only two pressure regulators are requ ired. A section of the flow-through reaction chamber is heated by ther moelectric modules to control the sequencing reaction temperature. Con version of the extracted amino acid product to the more stable phenylt hiohydantoin (PTH) form is performed off-line so that it may be dissol ved in 1 mu l buffer for identification. Approximately 0.1% of this PT H product is analyzed by micellar electrokinetic capillary chromatogra phy (MECC) with thermo-optical absorbance detection (TOAD), providing femtomole detection of the phenylthiohydantoin amino acids. Preliminar y results at the 50 pmol level for both adsorptive and covalent sequen cing methods are presented. (C) 1997 Elsevier Science B.V.