Deep-dive Targeted Quantification for Ultrasensitive Analysis of Proteins in Nondepleted Human Blood Plasma/Serum and Tissues
Nie S, Shi T, Fillmore TL, Schepmoes AA, Brewer H, Gao Y, Song E, Wang H, Rodland KD, Qian WJ, Smith RD, Liu T. Deep-Dive Targeted Quantification for Ultrasensitive Analysis of Proteins in Nondepleted Human Blood Plasma/Serum and Tissues. Anal Chem. 2017 Sep 5;89(17):9139-9146. doi: 10.1021/acs.analchem.7b01878. Epub 2017 Aug 11. PMID: 28724286; PMCID: PMC5627997
- Organism: Homo sapiens
- Instrument: TSQ Vantage,nanoACQUITY UPLC
- SpikeIn:
No
Mass spectrometry-based targeted proteomics (e.g., selected reaction monitoring, SRM) is emerging as an attractive alternative to immunoassays for protein quantification. Recently we have made significant progress in SRM sensitivity for enabling quantification of low ng/mL to sub-ng/mL level proteins in nondepleted human blood plasma/serum without affinity enrichment. However, precise quantification of extremely low abundant but biologically important proteins (e.g., ≤100 pg/mL in blood plasma/serum) using targeted proteomics approaches still remains challenging. To address this need, we have developed an antibody-independent Deep-Dive SRM (DD-SRM) approach that capitalizes on multidimensional high-resolution reversed-phase liquid chromatography (LC) separation for target peptide enrichment combined with precise selection of target peptide fractions of interest, significantly improving SRM sensitivity by ~5 orders of magnitude when compared to conventional LC-SRM. Application of DD-SRM to human serum and tissue has been demonstrated to enable precise quantification of endogenous proteins at ~10 pg/mL level in nondepleted serum and at <10 copies per cell level in tissue. Thus, DD-SRM holds great promise for precisely measuring extremely low abundance proteins or protein modifications, especially when high-quality antibody is not available.
Created on 5/3/17, 5:09 PM