title: Bead-Extractor Assisted Ready-to-Use Reagent System (BEARS) for Immunoprecipitation Coupled to MALDI-MS.
abstract: Quantitative protein assays play an important role in the study of biological functions. Immunoassays and mass spectrometry are two main technologies for quantifying proteins in biological samples. The combination of immunoprecipitation (IP) with MALDI technology delivers high assay sensitivity and specificity, but the sample preparation procedure involves multiple washing and transfer steps. These steps can be performed either manually (requiring significant time and labor) or automatically (requiring the purchase of a complex liquid-handling workstation). This bottleneck has limited the widespread adoption of this technology. We present here the Bead-Extractor Assisted ready-to-use Reagent System (BEARS) technology for simplified, low cost protein and peptide immunoprecipitation combined with MALDI-MS detection. All of the reagents are stable during long-term storage and can be prepared in advance. In the BEARS technology, a magnetic-bead extractor is used to handle beads from 96 wells simultaneously. A BEARS-based method was developed for plasma renin activity (PRA) and was evaluated on fifty-three clinical samples. These experiments showed that the BEARS assay had an LOD and linear range comparable to the manual method and an automated iMALDI PRA assay, but was 4-times faster than the manual approach. The BEARS iMALDI results also correlated well with a conventional ELISA PRA assay, with a coefficient of determination of 0.98. The BEARS technology provides convenience and affordability, and extends the use of IP-based mass spectrometry technology to most research and clinical laboratories, including those in developing countries.
authors.affiliation: University of Victoria - Genome BC Proteomics Centre, #3101-4464 Markham Street, Vancouver Island Technology Park, Victoria, British Columbia V8Z 7X8, Canada.|Biomedical Engineering Department, McGill University , Duff Medical Building, McGill University, 3775, rue University, Room 316, Montreal, Quebec H3A 2B4, Canada.|McGill University and Genome Quebec Innovation Centre , 740 Dr. Penfield Avenue, Room 6206, Montreal, Quebec H3A 1A4, Canada.
authors.forename: Huiyan
authors.initials: H
authors.lastname: Li
authors.affiliation: University of Victoria - Genome BC Proteomics Centre, #3101-4464 Markham Street, Vancouver Island Technology Park, Victoria, British Columbia V8Z 7X8, Canada.
authors.forename: Robert
authors.initials: R
authors.lastname: Popp
authors.affiliation: Jewish General Hospital , 3755 Côte-Ste-Catherine Road, Montreal, Quebec H3T 1E2, Canada.
authors.forename: Michael
authors.initials: M
authors.lastname: Chen
authors.affiliation: Jewish General Hospital , 3755 Côte-Ste-Catherine Road, Montreal, Quebec H3T 1E2, Canada.
authors.forename: Elizabeth M
authors.initials: EM
authors.lastname: MacNamara
authors.affiliation: Biomedical Engineering Department, McGill University , Duff Medical Building, McGill University, 3775, rue University, Room 316, Montreal, Quebec H3A 2B4, Canada.|McGill University and Genome Quebec Innovation Centre , 740 Dr. Penfield Avenue, Room 6206, Montreal, Quebec H3A 1A4, Canada.
authors.forename: David
authors.identifier: 0000-0002-7313-1162
authors.initials: D
authors.lastname: Juncker
authors.affiliation: University of Victoria - Genome BC Proteomics Centre, #3101-4464 Markham Street, Vancouver Island Technology Park, Victoria, British Columbia V8Z 7X8, Canada.|Department of Biochemistry and Microbiology, University of Victoria , Petch Building Room 207, 3800 Finnerty Road, Victoria, British Columbia V8P 5C2, Canada.|Gerald Bronfman Department of Oncology, Jewish General Hospital, McGill University , Montreal, Quebec, H3T 1E2, Canada.|Proteomics Centre, Segal Cancer Centre, Lady Davis Institute, Jewish General Hospital, McGill University , Montreal, Quebec, H3T 1E2, Canada.
authors.forename: Christoph H
authors.identifier: 0000-0003-2394-6512
authors.initials: CH
authors.lastname: Borchers