Automated sample preparation with SP3 for low-input clinical proteomics

ABSTRACT: High-throughput and streamlined workflows are essential in clinical proteomics for standardized processing of samples from a variety of sources, including fresh frozen tissue, FFPE tissue, or blood. To reach this goal, we have implemented single-pot solid-phase-enhanced sample preparation (SP3) on a liquid handling robot for automated processing (autoSP3) of tissue lysates in a 96-well format. AutoSP3 performs unbiased protein purification and digestion, and delivers peptides that can be directly analyzed by LCMS, thereby significantly reducing hands-on time, reducing variability in protein quantification, and improving longitudinal reproducibility. We demonstrate the distinguishing ability of autoSP3 to process low-input samples, reproducibly quantifying 500-1000 proteins from 100-1000 cells. Furthermore, we applied this approach to a cohort of clinical FFPE pulmonary adenocarcinoma (ADC) samples, and recapitulated their separation into known histological growth patterns. Finally, we integrated autoSP3 with AFA-ultrasonication for the automated end-to-end sample preparation and LCMS analysis of 96 intact tissue samples. Collectively this constitutes a generic, scalable, and cost-effective workflow with minimal manual intervention, enabling reproducible tissue proteomics in a broad range of clinical and non-clinical applications.

SUBMITTER: Mr Torsten Mueller 

PROVIDER: S-SCDT-MSB-19-9111 | biostudies-other |

REPOSITORIES: biostudies-other