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Single Cell Analysis Using Microfluidics Coupled to Ultrasensitive Mass Spectrometry


EMSL Project ID
47618

Abstract

Proteomics and metabolomics measurements in their present form require large populations of cells and thus average over and obscure important heterogeneity that is present even in clonal populations cultivated under highly controlled conditions [1]. For “real world” samples, this means that important but rare events go undetected, and the effects of stochastic expression and the microenvironment are blurred. The objective of this proposal is to combine microfluidic sample preparation and separations with the ultrasensitive mass spectrometry (MS) capability located in the EMSL to extend proteomic and metabolomic analyses to the level of single eukaryotic cells. The payoff will be significant, providing a unique capability for the EMSL in a field that is receiving greatly increased attention from the DOE [2], the NIH [3] and the scientific community at large. Nucleic acid analysis has matured rapidly for single cells [4], but the direct analysis of proteins and metabolites is significantly lacking. At present, the only established option is flow cytometry [5, 6], which is best suited to labeling cell surface markers in suspension cells and is limited to analytes having an available immunofluorescent tag. In contrast, MS provides for information-rich analyses in which chemical species can be unambiguously identified due to its high mass measurement accuracy and resolution. MS is also broad-band or multiplexed in that large numbers of analytes, including proteins, peptides and metabolites can be detected simultaneously without the need for chemical labeling. We will combine our expertise in the fields of microfluidics, chemical separations and ultrasensitive mass spectrometry to isolate, prepare and analyze hundreds of individual eukaryotic cells at an anticipated coverage of at least 50 proteins and metabolites per cell. We will use cultured mammalian cells for our initial tests, but the platform will be adaptable to a wide range of biological systems.

Project Details

Start Date
2012-09-04
End Date
2014-09-30
Status
Closed

Team

Principal Investigator

Ryan Kelly
Institution
Brigham Young University

Co-Investigator(s)

Bryan Linggi
Institution
Environmental Molecular Sciences Laboratory

Team Members

Levi Broeske
Institution
Pacific Northwest National Laboratory

Sarah Rausch
Institution
Environmental Molecular Sciences Laboratory

Andreas Vasdekis
Institution
University of Idaho

Xuefei Sun
Institution
Pacific Northwest National Laboratory

Keqi Tang
Institution
Pacific Northwest National Laboratory

Related Publications

Jambovane SR, SA Prost, AM Sheen, JK Magnuson, and RT Kelly. 2014. "ON-DEMAND SERIAL DILUTION USING QUANTIZED NANO/PICOLITER-SCALE DROPLETS." In 18th International Conference on Miniaturized Systems for Chemistry and Life Sciences, pp. 1247-1249. Chemical and Biological Microsystems Society, San Diego, CA.
Kelly RT, AM Sheen, and SR Jambovane. 2013. "Multilayer Microfluidic Devices Created From A Single Photomask." RSC Advances. [In Press]
Kelly RT, I Marginean, and K Tang. 2013. "Electrospray Ionization Mass Spectrometry." In Encyclopedia of Microfluidics and Nanofluidics. PNNL-SA-94681, Pacific Northwest National Laboratory, Richland, WA. [Unpublished]
Marginean I, K Tang, RD Smith, and RT Kelly. 2014. "Picoelectrospray Ionization Mass Spectrometry Using Narrow-bore Chemically Etched Emitters." Journal of the American Society for Mass Spectrometry 25(1):30-36. doi:10.1007/s13361-013-0749-z
Sun X, K Tang, RD Smith, and RT Kelly. 2013. "Controlled Dispensing and Mixing of Pico- to Nanoliter Volumes Using On-Demand Droplet-Based Microfluidics." Microfluidics and Nanofluidics 15(1):117-126. doi:10.1007/s10404-012-1133-1