Antibody-free, targeted mass-spectrometric approach for quantification of proteins at low picogram per milliliter levels in human plasma/serum
The detection and quantification of proteins at sub-ng/mL concentrations in plasma/serum is of critical importance for candidate biomarker verification. Sensitive detection of serum proteins has typically been achieved by immunoassays; however, de novo development of antibodies is associated with high cost and long lead time. To address this challenge, we developed an antibody-free strategy, termed high-pressure high-resolution separations with intelligent selection and multiplexing (PRISM), for achieving accurate detection of proteins at ~50 pg/mL level in plasma/serum using selected reaction monitoring (SRM). This strategy is based on high resolution reversed phase liquid chromatographic separations for analyte enrichment along with intelligent selection of target fractions via on-line SRM monitoring of internal standards and fraction multiplexing prior to SRM quantification. Our work represents a major technological advance for achieving pg/mL level of serum protein quantification without specific affinity reagents and holds great promise for broad applications in biomarker verification and systems biology studies.
- Research Organization:
- Pacific Northwest National Laboratory (PNNL), Richland, WA (US), Environmental Molecular Sciences Laboratory (EMSL)
- Sponsoring Organization:
- USDOE
- DOE Contract Number:
- AC05-76RL01830
- OSTI ID:
- 1059199
- Report Number(s):
- PNNL-SA-83776; 40072; 46206; 40991; 43796; 400412000
- Journal Information:
- Proceedings of the National Academy of Sciences of the United States of America, 109(38):15395-15400, Journal Name: Proceedings of the National Academy of Sciences of the United States of America, 109(38):15395-15400
- Country of Publication:
- United States
- Language:
- English
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