ASSESSING AUTOMATED SAMPLE PREPARATION TECHNOLOGIES FOR HIGH-THROUGHPUT PROTEOMICS OF FROZEN WELL CHARACTERIZED TISSUES FROM SWEDISH BIOBANKS

Large cohorts of carefully collected and stored clinical tissue materials play a central role in acquiring sufficient depth and statistical power to discover disease-related mechanisms and biomarkers of clinical significance. Manual preparation of such heterogeneous samples is labor intensive and requires experienced laboratory personnel. This carries other possible downsides such as low throughput, high risk of errors and low reproducibility, which limits the chances of making new Biomarker discoveries. In this work, three automated technologies for high-throughput proteomics of frozen sectioned tissues from our biobank were assessed. The instruments evaluated included the Bioruptor for tissue disruption and protein extraction; the Barocycler, which is able to disrupt tissues and digest the proteins; and the Bravo AssayMAP, a micro-chromatography platform for protein denaturation, digestion, peptide desalting and fractionation. A wide variety of tissue samples from 1) rat spleen, 2) human melanoma tumor, 3) human pancreatic tumor and 4) pancreatic tumor xenografts were assessed. The three instruments displayed reproducible and consistent results, as was proven by the high correlation between the measurements and by low coefficients of variation for the protein intensities for the different tissue types. Furthermore, we found that there is a good correlation between the obtained protein amount from the respective tumor tissues and the surface area of the analyzed tissue. This is of particular importance as the applicability is favored even in heterogeneous tumors originating form malignant melanoma patients. The results from this study has allowed us to integrate these technologies into an automated sample preparation workflow for large-scale proteomic studies that are currently ongoing.