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Combining microfluidic networks and peptide arrays for multi-enzyme assays

Su, Jing and Bringer, Michelle R. and Ismagilov, Rustem F. and Mrksich, Milan (2005) Combining microfluidic networks and peptide arrays for multi-enzyme assays. Journal of the American Chemical Society, 127 (20). pp. 7280-7281. ISSN 0002-7863. doi:10.1021/ja051371o. https://resolver.caltech.edu/CaltechAUTHORS:20130821-160733236

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Abstract

This paper reports the use of microfluidic networks (μFNs) to both prepare peptide microarrays and carry out label-free enzyme assays on self-assembled monolayers (SAMs) of alkanethiolates on gold. A poly(dimethylsiloxane) (PDMS) stamp fabricated with microchannels is used to immobilize a linear array of cysteine-terminated peptides onto SAMs presenting maleimide groups. The stamp is then reapplied to the SAM in a perpendicular direction to introduce enzyme solutions so that each solution can interact with an identical linear array of immobilized peptides. The μFNs enable multiple enzyme−substrate interactions to be simultaneously evaluated at a submicroliter scale, while the use of SAMs enables the use of MALDI mass spectrometry (MS) to analyze the enzyme activities. This paper demonstrates applications of this system for assaying multiple kinases and for profiling the activities of kinases and phosphatases in human K562 cell extracts. The combination of μFN, SAMs, and MS detection provides a flexible platform for assaying enzyme activities in biological samples.


Item Type:Article
Related URLs:
URLURL TypeDescription
http://dx.doi.org/10.1021/ja051371oDOIArticle
http://pubs.acs.org/doi/full/10.1021/ja051371oPublisherArticle
http://pubs.acs.org/doi/suppl/10.1021/ja051371oPublisherSupporting Information
ORCID:
AuthorORCID
Ismagilov, Rustem F.0000-0002-3680-4399
Additional Information:Copyright © 2005 American Chemical Society. Published In Issue: May 25, 2005. Received March 3, 2005. This work was supported by the NSF-MRSEC and used the microfluidics and protein expression facilities. Supporting Information Available: Details for fabrication of μFNs, preparation of SAMs, enzyme assays, mass spectrometric analysis, and cell extract preparation. This material is available free of charge via the Internet at http://pubs.acs.org.
Funders:
Funding AgencyGrant Number
NSFUNSPECIFIED
Subject Keywords:self-assembled monolayers; mass-spectrometry; selective inhibitor; tyrosine kinase; chip; poly(dimethylsiloxane); immunoassay; fabrication; devices; systems
Issue or Number:20
DOI:10.1021/ja051371o
Record Number:CaltechAUTHORS:20130821-160733236
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20130821-160733236
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:40877
Collection:CaltechAUTHORS
Deposited By: Whitney Barlow
Deposited On:28 Aug 2013 18:45
Last Modified:10 Nov 2021 04:23

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