Projects

Small Molecules Quantification

Our laboratory has developed targeted LC-MS/MS clinical and research assays for vitamin D and its metabolites, many different drugs in urine, and metanephrines in plasma. Our recent publication describing a testosterone assay for human serum samples demonstrated how important the technology can be for the appropriate care of patients. Our multiplexed assay for sphingolipids has helped elucidate the importance of these molecules in human health and our assay that quantifies multiple renally secreted molecules in urine and plasma has demonstrated the interesting complementarity that renal secretion has with glomerular filtration and protein retention in describing kidney disease.

Protein Quantification

We have helped pioneer the quantification of proteins in clinical samples. Our thyroglobulin assay that used peptide immunoaffinity enrichment-LC-MS/MS was the first proof-of-principle assay for a clinically relevant analyte described in the literature. We have also described assays for apoA-I/apoB, vitamin D binding protein, insulin-like growth factor-1, and retinol binding protein that helped lay the foundation for future advances in the technology. Our recent assays for apoE in CSF, biomarkers of collagen turnover in plasma, insulin/C-peptide in plasma, apoL1 in different sample types, and amyloid proteins in tissue have continued to push the field forward. Our current grant-funded efforts are focused on developing assays for type 1 diabetes, cancer, and Alzheimer's disease.

Biomarkers of type III collagen turnover

A novel method for the simultaneous quantification of peptides from the carboxy-terminal region of the type III procollagen, biomarkers of collagen turnover.
Current methods for identifying athletes who are using growth hormone for performance enhancement in sport include measuring the serum concentration of the amino-terminal propeptide liberated during the deposition of type III collagen (PIIINP). We have generated two novel monoclonal antibodies that can enrich peptides from other portions of the type III collagen molecule, including the carboxyl-terminal propeptide of type III procollagen (PIIICP) and breakdown fragments of the collagen fibril itself. Using these antibodies, we have developed a novel assay for collagen fragments in human plasma, which appear to change upon controlled administration of exogenous growth hormone.

Multi-Lab Research Program Studies Proteins in Alzheimer’s disease

Several research labs at UW SOM and Stanford University will be part of a new program that studies cerebrospinal fluid proteins in Alzheimer's & other dementias. This program, funded by a grant expected to total $15.9 million over five years from NIH, aims to apply recent advances to improve diagnosis, monitoring, and treatment. Our lab leads the development of laboratory tests for disease management. Other projects are led by the MacCoss Lab in the UW Department of Genome Sciences, Dr. Baker from the UW Medicine Institute of Protein Design, and Dr. Montine (former Chair of UW Pathology) from Stanford School of Medicine.

Multiplexed quantification of insulin and C-peptide by LC-MS/MS without the use of antibodies

The measurement of insulin and C-peptide provides a valuable tool for the clinical evaluation of hypoglycemia. In research, these biomarkers are used together to better understand hyperinsulinemia, hepatic insulin clearance, and beta cell function. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) is an attractive approach for the analysis of insulin and C-peptide because the platform is specific, can avoid certain limitations of immunoassays, and can be multiplexed. Previously described LC-MS/MS methods for the simultaneous quantification of insulin and C-peptide measure the intact analytes and most have relied on immunoaffinity enrichment. These approaches can be limited in terms of sensitivity and interference from auto-antibodies, respectively. We have developed a novel method that does not require antibodies and uses proteolytic digestion to yield readily ionizable proteotypic peptides that enables the sensitive, specific, and simultaneous quantitation of insulin and C-peptide.

Nutrition Obesity Research Center/Diabetes Research Center