STAGE-diging: a novel in-gel digestion processing for proteomics samples

frac1Autore del lavoro candidato: Paolo Soffientini

SINTESI CONTENENTE UNA BREVE DESCRIZIONE DEL LAVORO SVOLTO E DEI RISULTATI OTTENUTI: Proteomics is a very important, high throughput tool to identify ALL the proteins involved in a certain process. It requires the purification of all the proteins, their separation by gel electrophoresis and their digestion directly within the gel. The digested material is then processed by mass spectrometry that is able to identify all the proteins present. Proteomics is now actively researched in the clinical field as the presence of group of specific proteins may be extremely indicative of certain diseases, to response to therapies, hence to diagnostics and therapeutics. However, the complexity of the operations needed to prepare proteins for mass spectrometric analysis and the time required make it very difficult to adopt in clinical environments. We devised a new method, named STAGE-diging, for sample preparation in proteomics experiments, that can be easily adopted by any laboratory, and compared it with the standard in gel digestion procedure for sample preparation, both on high and low complexity protein samples. Moreover, we also tested the new method on quantitative proteomics experiments and on two different mass spectrometers. The results showed that STAGE-diging reduces sample handling, spare analysis time by more than 30% and improves protein identification and quantification (+13%). Moreover, it allows shorter instrument time and hence the possibility to perform multiple replicates that produce wider proteome coverage and more accurate protein quantitation. In addition, using this novel approach allows performing proteomics analyses on multiple samples, just as it would be necessary in the clinics. All these benefits make this new method appealing in case of large numbers of samples, where time and reproduci bility play a substantial role and enable the clinical application of the proteomics approach.