May 6, 2014 – MSDG Meeting

I. James Settlage Ph.D.

Inventiv Health Clinical, Princeton, NJ

“Leveraging the power of Supercritical Fluid Chromatography coupled with Triple Quadrupole Mass Spectrometry to meet otherwise intractable bioanalytical challenges”

II. Gene Ciccimaro, Ph.D.

Bristol-Myers Squibb, Lawrenceville, NJ

“Meeting the Needs of Biopharma Drug Discovery by Increasing Sensitivity Using ionKey Technology and Protein Immunoenrichment Cleanup”

Abstract for Talk II:

Current challenges facing the Discovery Bioanalyst include the need to understand the biology of preclinical disease models and demonstrate on target effects. Perturbations of biomarkers, protein ligands and receptors need to be quantified in both circulation and in tissues from target organs. Bioanalytical assays to provide this data require exquisite sensitivity due not only to extremely low endogenous levels, but also limited sample volume. To overcome these demands, we attempt to incorporate sample cleanup using immunoenrichment followed by low flow LC-MS technology using the ionKey micro column tile emitter and TQS mass spectrometer. There is a known asymptotic increase in analyte ESI-MS signal response with reduced solvent flow rate. Considering optimal QqQ instrumental ion transfer efficiency loss, a reduction from traditional flow rate (400-800 µL/min) to low micro-flow rates (1-5 µL/min) will on average result in 10-30X, and a reduction to nano-flow rate (<1 µL/ min) will result in ~ 50X gain in signal intensity. These intensity gains however, do not directly correlate to gains in analytical assay sensitivity (reduced LLOQ). In this study we highlight this disconnect when porting a protein immunoenrichment method from traditional to micro-flow LC-MS/MS. We identify charge competition as the cause for the performance gap, and show results from an improved sample preparation that achieves improved assay performance by maximizing sample loading and minimizing charge saturation. We highlight the use of this technology to support drug discovery efforts. Authors: Bogdan Sleczka, Eugene Ciccimaro, John Mehl, Lorell Discenza, Asoka Ranasinghe, Celia D’Arienzo, Timothy Olah (Bristol-Myers Squibb, Lawrenceville, NJ). Catalin Doneau, Brad Coopersmith, Jim Murphy (Waters)