Coupling microdroplet-based sample preparation, multiplexed isobaric labeling, and nanoflow peptide fractionation for deep proteome profiling of tissue microenvironment

ABSTRACT: Existing spatially-resolved proteomics technologies cannot provide deep proteome coverage due to limited sensitivity and poor sample recovery. Herein, we seamlessly combined laser capture microdissection with a low-volume sample processing technology that includes a microfluidic device named microPOTS (Microdroplet Processing in One pot for Trace Samples), multiplexed isobaric labelling, and a nanoflow peptide fractionation approach. The integrated workflow allowed us to maximize proteome coverage of laser-isolated tissue samples containing nanogram level of proteins. We demonstrated that the deep spatial proteomics platform can quantify more than 5,000 unique proteins from a small-sized human pancreatic tissue pixel (~60,000 um2) and differentiate unique protein abundance patterns in pancreas. Further, the use of microPOTS chip eliminated the requirement for advanced microfabrication capabilities and specialized nanoliter liquid handling equipment, making it more accessible to proteomic laboratories.

INSTRUMENT(S): Q Exactive HF-X

ORGANISM(S): Homo Sapiens (ncbitaxon:9606)

SUBMITTER: Paul D. Piehowski  

PROVIDER: MSV000093925 | MassIVE | Thu Jan 25 22:34:00 GMT 2024

SECONDARY ACCESSION(S): PXD048911

REPOSITORIES: MassIVE