Proteomics

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Multimodal measurement of the transcriptome and proteome from dissociated human pancreatic islet cells using nanoSPLITS

ABSTRACT: Human pancreatic islets from two human donors (Prodo Laboratories) were acquired and cultured for 24 hr in pancreatic islet media (PIMS, Prodo Laboratories). Single cells were dissociated with TrypLE for ~15-30 minutes, washed with PBS, stained with Calcein-AM/propidium iodide, and sorted onto nanoSPLITS chips using a cellenONE cell sorter. For the proteomic sample prep all steps utilized the nanoPOTS sample handling robot. Lysates were reduced with DTT, alkylated with IAA, digested overnight with trypsin/Lys-C at 37C, and vacuum dried / stored at -20C. LC-MS/MS data was acquired using TIFF DDA methodology on a Thermo Tribrid mass spectrometer. Data was searched using FragPipe version 20.0 before downstream analysis in R.

INSTRUMENT(S): Orbitrap Fusion Lumos

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

SUBMITTER: James M. Fulcher  

PROVIDER: MSV000093330 | MassIVE | Tue Nov 07 17:16:00 GMT 2023

REPOSITORIES: MassIVE

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Publications

Single-cell multiomics provides comprehensive insights into gene regulatory networks, cellular diversity, and temporal dynamics. Here, we introduce nanoSPLITS (nanodroplet SPlitting for Linked-multimodal Investigations of Trace Samples), an integrated platform that enables global profiling of the transcriptome and proteome from same single cells via RNA sequencing and mass spectrometry-based proteomics, respectively. Benchmarking of nanoSPLITS demonstrates high measurement precision with deep pr  ...[more]

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