Clinical Proteomics

Activities and Methods:

Dried Blood Spot (DBS) sampling for Multiple Reaction Monitoring Mass Spectrometry (MRM/MS)
The combination of DBS sampling with MRM-MS is the standard approach for small molecule biomarker analysis in nation-wide newborn screening centers. The ease of sample collection and storage has reduced costs and improved care for millions of patients per year. However, the application of DBS-MRM to protein biomarker analysis only recently been proposed. We are established leaders in advancing DBS-MRM towards clinical proteomic applications by reporting the first multiplexed DBS-MRM method for endogenous proteins in DBS samples (Chambers et al. Mol Cell Proteomics. 2013). Additionally, we have determined that most proteins in whole blood are also recoverable from DBS samples using untargeted MS approaches (Chambers et al. J Am Soc Mass Spectrom. 2013). More recently, we are developing a novel DBS-MRM assay for quantifying >90 protein targets in DBS samples and this shows considerable promise for the integration of DBS sampling into proteomics applications.

Immuno-Matrix Assisted Laser Desorption Mass Spectrometry (iMALDI)
iMALDI combines the sensitivity of immunoaffinity capture with the specificity of MS detection. In this novel approach, clinical samples (e.g. plasma, serum) are digested with trypsin and spiked with stable isotope-labeled standard (SIS) peptides as internal standards. The endogenous peptide and SIS peptide are co-captured by antibodies immobilized on magnetic beads and the entire bead-analyte complex is spotted directly on a MALDI target. Captured peptides are released from the antibody by the addition of an acidic MALDI matrix. We have now taken a multifaceted approach for translating our iMALDI technology into clinical laboratories for routine protein quantification. First, we have automated the sample preparation using the Agilent Bravo liquid handling robot for improved sample throughput. Secondly, we have optimized iMALDI assays for the Bruker Microflex MALDI-TOF, a bench-top instrument that is already widely used in regulated healthcare environments.

Results Highlights:

1) An Automated Assay for the Clinical Measurement of Plasma Renin Activity by immuno-MALDI (iMALDI). Popp R, Malmström D, Chambers AG, Lin D, Camenzind AG, van der Gugten JG, Holmes DT, Pugia M, Jaremek M, Cornett S, Suckau D, Borchers CH. Biochim Biophys Acta. 2014 Oct 16. pii: S1570-9639(14)00268-4. doi: 10.1016/j.bbapap.2014.10.008.

2) Comparison of proteins in whole blood and dried blood spot samples by LC/MS/MS. Chambers AG, Percy AJ, Hardie DB, Borchers CH. J Am Soc Mass Spectrom. 2013 Sep;24(9):1338-45. doi: 10.1007/s13361-013-0678-x.

3) Multiplexed quantitation of endogenous proteins in dried blood spots by multiple reaction monitoring-mass spectrometry. Chambers AG, Percy AJ, Yang J, Camenzind AG, Borchers CH. Mol Cell Proteomics. 2013 Mar;12(3):781-91. doi: 10.1074/mcp.M112.022442.

4) Development and evaluation of an immuno-MALDI (iMALDI) assay for angiotensin I and the diagnosis of secondary hypertension. Camenzind AG, van der Gugten JG, Popp R, Holmes DT, Borchers CH. Clin Proteomics. 2013 Dec 20;10(1):20. doi: 10.1186/1559-0275-10-20.

Andrew Chambers
Robert Popp
Juncong Yang
Darryl Hardie