While the development of new protein biomarkers is an exciting prospect, current proteomic measurement capabilities are lacking in key aspects, such as throughput, detection limits for low-abundance proteins, and data quality (including the confidence of protein identifications and under-sampling), and quantitation accuracy. In the result, the measurement quality is generally insufficient to confidently detect trace proteins in broad biomedical analyses, and the throughput possible does not allow the statistics of analyses needed for most purposes. The result is that measurement quality is generally insufficient to confidently detect trace proteins in broad analyses of biomedical samples, and the throughput possible is insufficient to provide statistically meaningful numbers of analyses for most purposes. In addition, most of the current proteomics platforms are best suited for detection of proteolytic peptides, using the "bottom-up" approach which is inefficient for distinguishing between biologically important protein isoforms and for identifications of post translational modifications of proteins. The overall objective of this project is to develop a platform for candidate protein biomarker discovery and verification analyses for application to human bodily fluids that will greatly improve upon existing methodologies in terms of throughput, sensitivity, robustness, and quantitation. The new platform aims at providing greater proteome coverage and enabling quantitative measurements of intact proteins and higher molecular weight peptides at concentrations that are presently problematic. The platform will encompass fast capillary zone electrophoresis (CZE) separation integrated with a novel microfabricated nano-electrospray ionization (nano-ESI) emitter on a microfluidic chip followed by a lower pressure field asymmetric ion mobility spectrometer (FAIMS) interfaced to a high resolution gas phase ion mobility (IMS) separation stage and a high-resolution time-of-flight mass spectrometer (TOF MS), providing accurate mass measurements.
The specific aims of this project are to: (1) design a single-use disposable microfluidic chip which incorporates CZE and nano-ESI source, (2) model and develop a reduced pressure FAIMS device for minimizing chemical background levels in analysis of intact proteins, (3) develop an efficient ion activation approach in conjunction with gas-phase IMS separation, and (4) rigorously evaluate the multidimensional proteomics platform with clinical samples. We anticipate that the proposed proteomics platform will provide the overall high sensitivity and separation power, as well as accurate quantitation, required for more effective and higher throughput measurements to discover assay patterns of proteins, protein fragments, and peptides in biological fluids that identify biological states.

Public Health Relevance

(provided by applicant): The specific objective of this project is to develop a high-throughput sensitive multidimensional separation platform for quantitative analysis of intact proteins and higher molecular weight peptides from human bodily fluids. The platform will enable condensed-phase protein separations and ionization using capillary zone electrophoresis and nano-electrospray ionization source on a single-use disposable microfluidic chip followed by efficient gas-phase ion mobility separations and mass spectrometry detection.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Exploratory/Developmental Grants (R21)
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Special Emphasis Panel (ZRR1-BT-7 (02))
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Friedman, Fred K
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Battelle Pacific Northwest Laboratories
United States
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Ibrahim, Yehia M; Garimella, Sandilya V B; Tolmachev, Aleksey V et al. (2014) Improving ion mobility measurement sensitivity by utilizing helium in an ion funnel trap. Anal Chem 86:5295-9
Prost, Spencer A; Crowell, Kevin L; Baker, Erin S et al. (2014) Detecting and removing data artifacts in Hadamard transform ion mobility-mass spectrometry measurements. J Am Soc Mass Spectrom 25:2020-7
Webb, Ian K; Garimella, Sandilya V B; Tolmachev, Aleksey V et al. (2014) Experimental evaluation and optimization of structures for lossless ion manipulations for ion mobility spectrometry with time-of-flight mass spectrometry. Anal Chem 86:9169-76
Garimella, Sandilya V B; Ibrahim, Yehia M; Webb, Ian K et al. (2014) Simulation of electric potentials and ion motion in planar electrode structures for lossless ion manipulations (SLIM). J Am Soc Mass Spectrom 25:1890-6
Shvartsburg, Alexandre A; Ibrahim, Yehia M; Smith, Richard D (2014) Differential ion mobility separations in up to 100% helium using microchips. J Am Soc Mass Spectrom 25:480-9
Webb, Ian K; Chen, Tsung-Chi; Danielson 3rd, William F et al. (2014) Implementation of dipolar resonant excitation for collision induced dissociation with ion mobility/time-of-flight MS. J Am Soc Mass Spectrom 25:563-71