| In recent years, ion mobility spectrometry (IMS) coupled to time-of-flight mass spectrometry (TOFMS) has become a powerful approach for understanding and characterizing complex biological biomolecules. The combined separation provides simultaneous measurement of an ion's size-to-charge and mass-to-charge ratios. When coupled to liquid chromatography (LC) separations, LC-IMS-TOFMS provides a high-throughput, multidimensional analytical platform with a high peak capacity for analyzing complex proteomic mixtures. In this approach, complex mixtures of proteins are enzymatically digested and the peptide mixture is separated by LC and analyzed by IMS-TOFMS. The IMS separation disperses the peptide ions prior to collisional activation at the exit of the drift tube, allowing sequencing information to be obtained for all ions in parallel, an advantage over scanning technologies. To exploit the advantages offered by IMS, improvements in this analytical technique are presented.;For characterization of proteomic mixtures, high-quality primary sequence information is necessary to positively identify the components with database search algorithms. To improve parallel sequencing capabilities with this technology, a new drift tube design has been developed. This design provides high-efficiency fragmentation and improves the transmission efficiency of the fragments to the time-of-flight mass spectrometer. Additionally, the new method of collisional activation utilizes a system of potential at the exit of the drift tube, allowing rapid modulation of the potentials to provide precursor and fragment ion information during the course of a single LC separation.;Tandem mass spectrometry has long provided sophisticated methods of characterizing complex systems. Inspired by these methods, we have developed a multidimensional ion mobility separation that allows ions of a specific mobility to be selected and activated between successive stages of IMS separation. Key to the development of this technology has been the incorporation of ion funnels at strategic locations within the drift tube that radially focus the diffuse ion beam to the center of the drift tube. Prior to this development, high-resolution, low-pressure ion mobility separations were not feasible due to low ion transmission. Current designs in our laboratory now extend to several meters in length and offer resolutions as high as 200. |
