Temporal Sampling of Enzymes from Live Cells by Localized Electroporation and Quantification of Activity by SAMDI Mass Spectrometry
Prithvijit Mukherjee
Department of Mechanical Engineering, Northwestern University, Evanston, IL, 60208 USA
Theoretical and Applied Mechanics Program, Northwestern University, Evanston, IL, 60208 USA
Search for more papers by this authorEric J. Berns
Department of Biomedical Engineering, Northwestern University, Evanston, IL, 60208 USA
Search for more papers by this authorCesar A. Patino
Department of Mechanical Engineering, Northwestern University, Evanston, IL, 60208 USA
Search for more papers by this authorElamar Hakim Moully
Department of Chemistry, Northwestern University, Evanston, IL, 60208 USA
Search for more papers by this authorLingqian Chang
Department of Mechanical Engineering, Northwestern University, Evanston, IL, 60208 USA
Search for more papers by this authorS. Shiva P. Nathamgari
Department of Mechanical Engineering, Northwestern University, Evanston, IL, 60208 USA
Theoretical and Applied Mechanics Program, Northwestern University, Evanston, IL, 60208 USA
Search for more papers by this authorJohn A. Kessler
Department of Neurology, Northwestern University Feinberg School of Medicine, Chicago, IL, 60611 USA
Search for more papers by this authorCorresponding Author
Milan Mrksich
Department of Biomedical Engineering, Northwestern University, Evanston, IL, 60208 USA
Department of Chemistry, Northwestern University, Evanston, IL, 60208 USA
Department of Cell and Development Biology, Northwestern University Feinberg School of Medicine, Chicago, IL, 60611 USA
E-mail: [email protected], [email protected]
Search for more papers by this authorCorresponding Author
Horacio D. Espinosa
Department of Mechanical Engineering, Northwestern University, Evanston, IL, 60208 USA
Theoretical and Applied Mechanics Program, Northwestern University, Evanston, IL, 60208 USA
E-mail: [email protected], [email protected]
Search for more papers by this authorPrithvijit Mukherjee
Department of Mechanical Engineering, Northwestern University, Evanston, IL, 60208 USA
Theoretical and Applied Mechanics Program, Northwestern University, Evanston, IL, 60208 USA
Search for more papers by this authorEric J. Berns
Department of Biomedical Engineering, Northwestern University, Evanston, IL, 60208 USA
Search for more papers by this authorCesar A. Patino
Department of Mechanical Engineering, Northwestern University, Evanston, IL, 60208 USA
Search for more papers by this authorElamar Hakim Moully
Department of Chemistry, Northwestern University, Evanston, IL, 60208 USA
Search for more papers by this authorLingqian Chang
Department of Mechanical Engineering, Northwestern University, Evanston, IL, 60208 USA
Search for more papers by this authorS. Shiva P. Nathamgari
Department of Mechanical Engineering, Northwestern University, Evanston, IL, 60208 USA
Theoretical and Applied Mechanics Program, Northwestern University, Evanston, IL, 60208 USA
Search for more papers by this authorJohn A. Kessler
Department of Neurology, Northwestern University Feinberg School of Medicine, Chicago, IL, 60611 USA
Search for more papers by this authorCorresponding Author
Milan Mrksich
Department of Biomedical Engineering, Northwestern University, Evanston, IL, 60208 USA
Department of Chemistry, Northwestern University, Evanston, IL, 60208 USA
Department of Cell and Development Biology, Northwestern University Feinberg School of Medicine, Chicago, IL, 60611 USA
E-mail: [email protected], [email protected]
Search for more papers by this authorCorresponding Author
Horacio D. Espinosa
Department of Mechanical Engineering, Northwestern University, Evanston, IL, 60208 USA
Theoretical and Applied Mechanics Program, Northwestern University, Evanston, IL, 60208 USA
E-mail: [email protected], [email protected]
Search for more papers by this authorAbstract
Measuring changes in enzymatic activity over time from small numbers of cells remains a significant technical challenge. In this work, a method for sampling the cytoplasm of cells is introduced to extract enzymes and measure their activity at multiple time points. A microfluidic device, termed the live cell analysis device (LCAD), is designed, where cells are cultured in microwell arrays fabricated on polymer membranes containing nanochannels. Localized electroporation of the cells opens transient pores in the cell membrane at the interface with the nanochannels, enabling extraction of enzymes into nanoliter-volume chambers. In the extraction chambers, the enzymes modify immobilized substrates, and their activity is quantified by self-assembled monolayers for matrix-assisted laser desorption/ionization (SAMDI) mass spectrometry. By employing the LCAD-SAMDI platform, protein delivery into cells is demonstrated. Next, it is shown that enzymes can be extracted, and their activity measured without a loss in viability. Lastly, cells are sampled at multiple time points to study changes in phosphatase activity in response to oxidation by hydrogen peroxide. With this unique sampling device and label-free assay format, the LCAD with SAMDI enables a powerful new method for monitoring the dynamics of cellular activity from small populations of cells.
Conflict of Interest
H.D.E. is the founder and majority owner of Infinitesimal LLC, a company commercializing bio-tools for gene editing and cell analysis. M.M. is the founder and chairman of SAMDI Tech Inc., which uses SAMDI-MS to assist clients in the pharmaceutical industry.
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