ESI/ion trap/ion mobility/time-of-flight mass spectrometry for rapid and sensitive analysis of biomolecular mixtures

An ion trap/ion mobility/time-of-flight mass spectrometry technique is show n to be a rapid and sensitive means of analyzing peptide/protein mixtures. In this approach, an ion trap is used to accumulate ions that have been ele ctrosprayed from a mixture into concentrated packets. The ion packets are i njected into a drift tube where components of the mixture are separated bas ed on differences in mobility through a buffer gas. Ions that exit the drif t tube are dispersed in a time-of-flight mass spectrometer for mass-to-char ge (m/z) determination. The gas-phase separation strategy reduces congestio n in the mass spectrum, and experimental mobilities complement m/z measurem ents in assigning peaks. Examples of the application of the approach to ide ntification of peptides (from tryptic digests) and to separation of charge- state distributions from electrospray of a mixture containing ubiquitin and myoglobin are presented. Most peptides that are observed from tryptic dige sts of proteins such as cytochrome c and myoglobin can be identified from d ata that are acquired in under 1 min; studies of mixtures with known compos itions indicate that detection limits are similar to 0.5-3 pmol for individ ual components. Factors that may influence the distributions that are obser ved, such as storage time in the trap, injection voltages used for the mobi lity experiment, and variations in ion cross section with charge state, are discussed.