Detecting Kinase Activities from Single Cell Lysate Using Concentration-Enhanced Mobility Shift Assay
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Han_Detecting Kinase.pdf
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Author(s)
Cheow, Lih Feng
Sarkar, Aniruddh
Kolitz, Sarah
Han, Jongyoon
Lauffenburger, Douglas A.
Date Issued
July 2014
Journal
Analytical Chemistry
Publisher
American Chemical Society (ACS)
Citation
Cheow, Lih Feng, Aniruddh Sarkar, Sarah Kolitz, Douglas Lauffenburger, and Jongyoon Han. “Detecting Kinase Activities from Single Cell Lysate Using Concentration-Enhanced Mobility Shift Assay.” Anal. Chem. 86, no. 15 (August 5, 2014): 7455–7462.
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Author's final manuscript
Abstract
Electrokinetic preconcentration coupled with mobility shift assays can give rise to very high detection sensitivities. We describe a microfluidic device that utilizes this principle to detect cellular kinase activities by simultaneously concentrating and separating substrate peptides with different phosphorylation states. This platform is capable of reliably measuring kinase activities of single adherent cells cultured in nanoliter volume microwells. We also describe a novel method utilizing spacer peptides that significantly increase separation resolution while maintaining high concentration factors in this device. Thus, multiplexed kinase measurements can be implemented with single cell sensitivity. Multiple kinase activity profiling from single cell lysate could potentially allow us to study heterogeneous activation of signaling pathways that can lead to multiple cell fates.
MIT Department
Massachusetts Institute of Technology. Department of Biological Engineering
Massachusetts Institute of Technology. Department of Electrical Engineering and Computer Science
Massachusetts Institute of Technology. Research Laboratory of Electronics
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Article is made available in accordance with the publisher's policy and may be subject to US copyright law. Please refer to the publisher's site for terms of use.
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DOI of Published Version
https://doi.org/10.1021/ac502185v
