High-throughput sample preparation for protein or peptide structural characterization

High-performance liquid chromatography (HPLC) combined with mass spectrometry has become the method of choice for identifying and characterizing cell-expressed biomolecules. This technology has evolved so rapidly that efficient sample preparation in a high-throughput mode has become a rate-limiting step. Applications using C18 resin, 200 A pore size, and 15-microm bead size silica and sulfonated divinylbenzene particles were tested. The convenient, solvent-resistant, 96-well MultiScreen filter plates with the Multiscreen Column Loader provided efficient removal of salts and detergents and excellent sample recovery for small volumes. The 96-well simultaneous, uniform loading of dry powders, beads, or resins in 45-, 80-, or 100-microL volumes easily accommodates various media capacities and elution volumes. Recovered eluates demonstrated high well-to-well reproducibility during analyte adsorption, washing, and elution. Sample recovery was analyzed by HPLC for a variety of proteins, peptides, and proteolytic digests. The utility of MultiScreen 96-well mini-columns in performing sample clean-up was also demonstrated for HPLC and mass spectroscopy.