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Michael Volny

Affiliate Scientist

Email

volny@apl.washington.edu

Publications

2000-present and while at APL-UW

High-throughput workflow for identification of phosphorylated peptides by LC-MALDI-TOF/TOF-MS coupled to in situ enrichment on MALDI plates functionalized by ion landing

Krásný, L., P. Pompach, M. Strnadová, R. Hynek, K. Vališ, V. Havlíček, P. Novák, and M. Volný, "High-throughput workflow for identification of phosphorylated peptides by LC-MALDI-TOF/TOF-MS coupled to in situ enrichment on MALDI plates functionalized by ion landing," J. Mass Spec., 50, 802-811, doi:10.1002/jms.3586, 2015.

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1 Jun 2015

We report an MS-based workflow for identification of phosphorylated peptides from trypsinized protein mixtures and cell lysates that is suitable for high-throughput sample analysis. The workflow is based on an in situ enrichment on matrix-assisted laser desorption/ionization (MALDI) plates that were functionalized by TiO2 using automated ion landing apparatus that can operate unsupervised. The MALDI plate can be functionalized by TiO2 into any array of predefined geometry (here, 96 positions for samples and 24 for mass calibration standards) made compatible with a standard MALDI spotter and coupled with high-performance liquid chromatography. The in situ MALDI plate enrichment was compared with a standard precolumn-based separation and achieved comparable or better results than the standard method. The performance of this new workflow was demonstrated on a model mixture of proteins as well as on Jurkat cells lysates. The method showed improved signal-to-noise ratio in a single MS spectrum, which resulted in better identification by MS/MS and a subsequent database search. Using the workflow, we also found specific phosphorylations in Jurkat cells that were nonspecifically activated by phorbol 12-myristate 13-acetate. These phosphorylations concerned the mitogen-activated protein kinase/extracellular signal-regulated kinase signaling pathway and its targets and were in agreement with the current knowledge of this signaling cascade. Control sample of non-activated cells was devoid of these phosphorylations. Overall, the presented analytical workflow is able to detect dynamic phosphorylation events in minimally processed mammalian cells while using only a short high-performance liquid chromatography gradient.

Nanoliter segmented-flow sampling mass spectrometry with online compartmentalization

Volný, M., J. Rolfs, B. Hakimi, P. Fryčák, T. Schneider, D. Liu, G. Yen, D.T. Chiu, and F. Tureček, "Nanoliter segmented-flow sampling mass spectrometry with online compartmentalization," Anal. Chem., 86, 3647-3652, doi:10.1021/ac500365r, 2014.

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13 Mar 2014

We report a microfluidic device, using segmented flow in a two-phase system of immiscible liquids, which delivers aqueous droplets into a modified commercial mass spectrometer. The interface coupling the microfluidics to the mass spectrometer achieves up to 96% sample transfer efficiency to the vacuum chamber. Sample ionization is assisted by multipass infrared laser beam in the interface. The system achieves low femtomole detection limits of several analytes ranging from drugs to proteins. Sample ionization in this segmented-flow sampling was found to be remarkably insensitive to the presence of buffer salts and other matrices.

Inventions

Reactive landing for modification of graphene and other nanocarbon materials

Record of Invention Number: 46967

Tim Elam, Joe Rolfs, Frantisek Turecek, Michael Volny

Disclosure

27 May 2014

Reactive Landing Resistance Biosensors

Record of Invention Number: 45652

Tim Elam, Frantisek Turecek, Michael Volny

Disclosure

8 Jun 2011

Acoustics Air-Sea Interaction & Remote Sensing Center for Environmental & Information Systems Center for Industrial & Medical Ultrasound Electronic & Photonic Systems Ocean Engineering Ocean Physics Polar Science Center
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