Reference: Tian R, et al. (2011) Development of a multiplexed microfluidic proteomic reactor and its application for studying protein-protein interactions. Anal Chem 83(11):4095-102

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Abstract


Mass spectrometry-based proteomics techniques have been very successful for the identification and study of protein-protein interactions. Typically, immunopurification of protein complexes is conducted, followed by protein separation by gel electrophoresis and in-gel protein digestion, and finally, mass spectrometry is performed to identify the interacting partners. However, the manual processing of the samples is time-consuming and error-prone. Here, we developed a polymer-based microfluidic proteomic reactor aimed at the parallel analysis of minute amounts of protein samples obtained from immunoprecipitation. The design of the proteomic reactor allows for the simultaneous processing of multiple samples on the same devices. Each proteomic reactor on the device consists of SCX beads packed and restricted into a 1 cm microchannel by two integrated pillar frits. The device is fabricated using a combination of low-cost hard cyclic olefin copolymer thermoplastic and elastomeric thermoplastic materials (styrene/(ethylene/butylenes)/styrene) using rapid hot-embossing replication techniques with a polymer-based stamp. Three immunopurified protein samples are simultaneously captured, reduced, alkylated, and digested on the device within 2-3 h instead of the days required for the conventional protein-protein interaction studies. The limit of detection of the microfluidic proteomic reactor was shown to be lower than 2 ng of protein. Furthermore, the application of the microfluidic proteomic reactor was demonstrated for the simultaneous processing of the interactome of the histone variant Htz1 in wild-type yeast and in a swr1? yeast strain compared to an untagged control using a novel three-channel microfluidic proteomic reactor.

Reference Type
Journal Article
Authors
Tian R, Hoa XD, Lambert JP, Pezacki JP, Veres T, Figeys D
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Interaction Annotations


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Interactor Interactor Type Assay Annotation Action Modification Phenotype Source Reference

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Gene Gene Ontology Term Qualifier Aspect Method Evidence Source Assigned On Annotation Extension Reference

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