The development of true microscale interfaces for protein mass spectrometry based on silicon chip micromachining is proposed. The devices would integrate electrospray ionization with a variety of protein isolation, digestion, and/or separation steps into a single monolithic structure that can be mass produced in much the same way as computer chips are made. The silicon chip devices would be coupled to state-of-the-art tandem mass spectrometer systems capable of providing detailed structural information on protein samples. Functional utility of the systems will be increased by the development of automated data collection procedures capable of analyzing incoming spectra and adjusting analysis parameters in real time. In addition to optimizing general factors such as relative collison energy and resolution, spectra would be evaluated in the context of the problem to be solved. A fast, multiprocessor computer will provide realtime access to a knowledge base and spectra will be analyzed according to a user defined set of rules. This expert system approach will be applied to a number of different kinds of applications including the rapid determination of protein variants and posttranslational modifications, the definition of specific protein protein interactions, and the identification of MHC complex peptides. A fundamental goal of the proposed research is to create a system in which both hardware and analytical expertise can be readily transferred to other labs and other researchers in the form of inexpensive, disposable interfaces and problem specific data collection and analysis programs. The development of such tools will greatly facilitate work to define the human proteome.
