Single Cell Proteomics with Ultra-High Sensitivity Mass Spectrometry

Frank, M

Lawrence Livermore National Laboratory

关键词: Desorption;    Ion Sources;    Velocity;    Aerosols;    Spectra;   

DOI  :  10.2172/15011526
RP-ID  :  UCRL-TR-210153
RP-ID  :  W-7405-ENG-48
RP-ID  :  15011526

美国|英语

来源: UNT Digital Library

【 摘 要 】

This project was a joint LDRD project between PAT, CMS and NAI with the objective to develop an instrument that analyzes the biochemical composition of single cells in real-time using bioaerosol mass spectrometry (BAMS) combined with advanced laser desorption and ionization techniques. Applications include both biological defense, fundamental cell biology and biomedical research. BAMS analyzes the biochemical composition of single, micrometer-sized particles (such as bacterial cells or spores) that can be directly sampled from air or a suspension. BAMS is based on an earlier development of aerosol time of flight mass spectrometry (ATOFMS) by members of our collaboration [1,2]. Briefly, in ATOFMS and BAMS aerosol particles are sucked directly from the atmosphere into vacuum through a series of small orifices. As the particles approach the ion source region of the mass spectrometer, they cross and scatter light from two CW laser beams separated by a known distance. The timing of the two bursts of scattered light created by each ''tracked'' particle reveals the speed, location and size of the particle. This information then enables the firing of a high-intensity laser such that the resulting laser pulse desorbs and ionizes molecules from the tracked particle just as it reaches the center of the ion source region. The full spectrum of ions is then measured using a time-of-flight mass spectrometer. The ability to rapidly analyze individual particles is clearly applicable to the rapid detection of aerosolized biological warfare agents so long as agent particles can be made to produce mass spectra that are distinct from the spectra of harmless background particles. The pattern of ions formed is determined by the properties of the laser pulse, the particle, and, in aerosol matrix-assisted laser desorption/ionization (MALDI), also the MALDI matrix used. As a result, it is critical that the properties of the laser pulses used for desorption and ionization be carefully chosen. The work on this LDRD project was centered on demonstrating the usefulness of mass signatures obtained by BAMS for identification of biological agents and discrimination from background. To accomplish this goal this work also included a thorough study of the origins of the observed mass signatures (i.e. peak identification and dependence on bacterial growth conditions) and their dependence on laser parameters. Such a fundamental understanding of the mass signatures and their dependence on laser parameters is required for optimizing the desorption/ionization process and the bioaerosol mass spectrometer in order to increase the sensitivity and specificity of this method for practical CBNP applications.