ポスター発表
- 第3日 5月17日（木） ポスター会場
-
3P-11 PDF
Extending quantitative proteome coverage through integrate data dependent and independent acquisition strategies
Data dependent acquisition (DDA) and the recently advancing data independent acquisition (DIA) MS strategies have both resulted in improved understanding of complexity of proteomics to study biological systems. In this study we aimed to achieve in depth proteome profiling through optimized single shot DDA and DIA. For targeted data extraction, we constructed reference spectral library of nearly 10,500 proteins (~183,000 peptides) from 98 LC-MS/MS DDA files acquired from HeLa cell lysate, non-small lung cancer cell lines (NSCLC) cell lines and pooled tumor tissue by single-shot and high pH reverse phase (HpRP) fractionation. Then we further evaluated several DIA acquisition parameters including precursor isolation window, precursor m/z ranges, MS and MS/MS resolution, injection time and gradient time. We were then able to quantify over 7300 proteins of which ~6300 were within 20% coefficient of variation (CV) from triplicate 160 min DIA acquisition resulting in almost two fold that of DDA. DIA quantitation against reference library provide reproducible deep quantitation. Finally, we were able to extend proteome spectral library by DDA and achieve improved DIA identification coverage and quantitation accuracy applicable to NSCLC cell lines proteome profiling.