A capillary monolithic trypsin reactor for efficient protein digestion in online and offline coupling to ESI and MALDI mass spectrometry

Analytical Chemistry
Jens Sproß, Andrea Sinz

Abstract

We describe the preparation of a capillary trypsin immobilized monolithic enzyme reactor (IMER) for a rapid and efficient digestion of proteins down to the femtomole level. Trypsin was immobilized on a poly(glycidyl methacrylate-co-acrylamide-co-ethylene glycol dimethycrylate) monolith using the glutaraldehyde technique. Digestion efficiencies of the IMER were evaluated using model proteins and protein mixtures as well as chemically cross-linked lysozyme regarding the addition of denaturants and increasing digestion temperature. The trypsin IMER described herein is applicable for the digestion of protein mixtures. Even at a 1000-fold molar excess of one protein, low-abundance proteins are readily identified, in combination with MS/MS analysis. An online setup of the IMER with reversed phase nano-HPLC separation and nano-ESI-MS/MS analysis was established. The great potential of the trypsin IMER for proteomics applications comprise short digestion times in the range of seconds to minutes, in addition to improved digestion efficiencies, compared to in-solution digestion.

References

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Aug 12, 2009·Analytical and Bioanalytical Chemistry·Jens Spross, Andrea Sinz

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Citations

Jan 21, 2012·Analytical and Bioanalytical Chemistry·Jens Spross, Andrea Sinz
May 31, 2012·Analytical and Bioanalytical Chemistry·Jens SproßAndrea Sinz
Jul 14, 2010·Amino Acids·Bjoern MeyerMichael Karas
Nov 21, 2014·Bioanalysis·Fred E Regnier, JinHee Kim
Mar 6, 2012·Journal of Pharmaceutical and Biomedical Analysis·E CalleriG Massolini
Aug 10, 2011·The FEBS Journal·Hu ZhouDaniel Figeys
Dec 18, 2013·Journal of Mass Spectrometry : JMS·Muhammad SafdarJanne Jänis
May 24, 2015·Annual Review of Analytical Chemistry·Jinlan Dong, Merlin L Bruening
Aug 26, 2014·Journal of Separation Science·Tingting HongYibing Ji
Jul 28, 2011·Journal of Separation Science·Jens Sproß, Andrea Sinz
Jan 30, 2020·Chembiochem : a European Journal of Chemical Biology·Jens SproßHarald Gröger
May 7, 2020·Journal of Materials Chemistry. B, Materials for Biology and Medicine·Chao ZhongZian Lin
Mar 16, 2013·Journal of Separation Science·Evgenia G Vlakh, Tatiana B Tennikova
Jan 8, 2013·Journal of Separation Science·Evgenia G Vlakh, Tatiana B Tennikova
Aug 28, 2020·Frontiers in Bioengineering and Biotechnology·Andoni Rodriguez-AbetxukoAna Beloqui
May 7, 2013·Journal of Materials Chemistry. B, Materials for Biology and Medicine·Ying ShenLanqun Mao
Feb 2, 2013·Journal of Proteome Research·Linda SwitzarWilfried M A Niessen
Apr 16, 2011·Analytical Chemistry·Zhibin NingDaniel Figeys

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