Multicentric study of the effect of pre-analytical variables in the quality of plasma samples stored in biobanks using different complementary proteomic methods.

Analytical proteomics has experienced exponential progress in the last decade and can be expected to lead research studies on diagnostic and therapeutic biomarkers in the near future. Because the development of this type of analysis requires the use of a large number of human samples with a minimum of quality requirements, our objective was to identify appropriate indicators for quality control of plasma samples stored in biobanks for research in proteomics. To accomplish this, plasma samples from 100 healthy donors were obtained and processed according to the pre-analytical variables of: a) time delay for the first centrifugation of the original blood sample (4 or 24h) and b) number of freeze/thaw cycles (1, 2 or 3) of the processed plasma samples. The analyses of samples were performed by different and complementary methods such as SPE MALDI-TOF, DIGE, shotgun (iTRAQ, nLC MALDI TOF/TOF) and targeted nLC MS/MS proteomic techniques (SRM). In general, because the distribution of proteins in all samples was found to be very similar, the results shown that delayed processing of blood samples and the number of freeze/thaw cycles has little or no effect on the integrity of proteins in the plasma samples. The results of the present work indicate that blood proteins in plasma are broadly insensitive to such preanalytical variables as delayed processing or freeze/thaw cycles when analyzed at the peptide level. Although there are other studies related to protein stability of clinical samples with similar results, what is remarkable about our work is the large number of plasma samples examined and that our analyses assessed protein integrity by combining a wide set of complementary proteomic approaches performed at different proteomic platform participating laboratories that all yielded similar results. We believe our study is the most comprehensive performed to date to determine the changes in proteins induced by delayed sample processing and plasma freeze/thaw cycles