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On-chip AP-MS: Interaction proteomics on a microfluidics platform

ABSTRACT: Here we present a microfluidics-based affinity purification-mass spectrometry (AP-MS) pipeline to identify protein-protein interactions (PPI) using minute amounts of input material. The use of this automated platform allowed us to identify the human cohesin and CCC complex from only 4 micrograms of input lysate, representing a ~50-100 fold downscaling compared to regular microcentrifuge tubes-based workflows. As such, our method holds great promise for future AP-MS applications in which sample amounts are limited.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Hela Cell

SUBMITTER: Cristina Furlan

LAB HEAD: Michiel Vermeulen

PROVIDER: PXD011323 | Pride | 2022-03-10

REPOSITORIES: Pride

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			Action	DRS
	1.5M_300K_60K_RESULTS.rar	Other
	1.5m_300k_60k_SMC1A_PD_4ug.rar	Other
	12K_Ab6556_SMC1A_RESULTS.rar	Other
	12kSC_BUBR1_mit_PD.rar	Other
	12kSC_BUBR1_mit_PD_RESULTS.rar	Other

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On-chip AP-MS: Interaction proteomics on a microfluidics platform

Project description:Essentially all cellular processes are orchestrated by protein-protein interactions (PPIs). In recent years, affinity purification coupled to mass spectrometry (AP-MS) has been the preferred method to identify cellular PPIs. Here we present a microfluidics-based (AP-MS) workflow to identify PPIs using minute amounts of input material. The use of this automated platform allowed us to identify the human cohesin, CCC and Mediator complex from as little as 4 micrograms of input lysate, representing a 50─100-fold downscaling compared to regular microcentrifuge tube-based protocols. We show that our platform can be used to affinity purify tagged baits as well as native cellular proteins and their interaction partners. As such, our method holds great promise for future biological and clinical AP-MS applications in which sample amounts are limited.

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