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Mechanistic investigation of the on surface enzymatic digestion (oSED) protein adsorption detection method using targeted mass spectrometry
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Uppsala University, Science for Life Laboratory, SciLifeLab.ORCID iD: 0000-0003-3861-9338
Uppsala University, Disciplinary Domain of Science and Technology, Technology, Department of Engineering Sciences, Microsystems Technology.
Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
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2016 (English)In: The Analyst, ISSN 0003-2654, E-ISSN 1364-5528, Vol. 141, no 5, 1714-1720 p.Article in journal (Refereed) Published
Abstract [en]

This study describes our efforts to study some of the mechanistic aspects of the earlier established onsurface enzymatic digestion (oSED) method. In a multitude of application areas, it has become important to be able to fully characterize and understand selective protein adsorption to biomaterial surfaces for various applications, including biomedicine (implants), nanotechnology (microchip surfaces and sensors) and materials sciences. Herein, the investigation of the mechanistic aspects was based on microdialysis catheter tubes that were flushed with controlled protein solutions mimicking the extracellular fluid of the brain. The protein adsorption properties were monitored using high-resolution liquid chromatography tandem mass spectrometry (LC-MS/MS) with a targeted method. The temporally resolved results show that most proteins stay adsorbed onto the surface during the entire digestion process and are only cut away piece by piece, whereas smaller proteins and peptides seem to desorb rather easily from the surface. This information will simplify the interpretation of data generated using the oSED method and can also be used for the characterization of the physicochemical properties controlling the adsorption of individual proteins to specific surfaces.

Place, publisher, year, edition, pages
2016. Vol. 141, no 5, 1714-1720 p.
National Category
Analytical Chemistry Engineering and Technology
Research subject
Chemistry with specialization in Analytical Chemistry
Identifiers
URN: urn:nbn:se:uu:diva-248629DOI: 10.1039/c5an02091cISI: 000371229600018OAI: oai:DiVA.org:uu-248629DiVA: diva2:800460
Funder
Magnus Bergvall FoundationBerzelii Centre EXSELENTSwedish Research Council, 621-2011-4423
Available from: 2015-04-05 Created: 2015-04-05 Last updated: 2017-12-04Bibliographically approved
In thesis
1. Characterization of Molecular Adsorption Using Liquid Chromatography and Mass Spectrometry
Open this publication in new window or tab >>Characterization of Molecular Adsorption Using Liquid Chromatography and Mass Spectrometry
2015 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Molecular adsorption is a key feature in several disciplines of chemistry, covering as diverse fields as chromatographic separation to biomaterial development. This thesis aims at developing methods and techniques for the characterization of molecular adsorption at the liquid-solid interface. Two different experimental models were used, small molecular interaction characterization using liquid chromatography and complex protein adsorption on polymeric materials possible for biological sampling. Holistic approaches, where both detailed molecular interactions and identifications of trends, could improve the fundamental understanding of adsorption systems, were invariably part of the scientific process.

The characterization of small molecular interactions on liquid chromatography stationary phases via adsorption isotherm determination used combined data from physical phase parameters i.e. carbon loading, linear-, and nonlinear-characterization methods. These experiments were conducted on high performance liquid chromatography systems, using both ordinary reversed phase stationary phases, and hybrid phases. The expansion of the improved elusion by characteristic point (ECP) for adsorption isotherm determination, led to that previous impossible isotherm types, having inflexion points, now could be determined by the method. It also reduced errors in isotherm parameters due to the elimination of inaccurate determined retention times where the mobile phase concentration was zero.

The characterization of protein adsorption where performed in an unbiased way. Adsorbed proteins on different surfaces were identified using mass spectrometry (MS) and data dependent acquisition or a targeted method. Prior MS, an improved on surface enzymatic digestion (oSED) method was used to enable identification and quantitation of adsorbed protein originating from ventricular cerebrospinal fluid (vCSF). oSED was found to be able to experimentally determine large variations in protein adsorption characteristics between native and coated polycarbonate surfaces in contact with vCSF. The method was also confirmed being mechanistic in favor of enzymatic digestion of the proteins adsorbed on a surface, rather than a prior desorption into solution before digestion.

An improvement of the overall understanding of adsorption systems was not only achieved with the oSED method as a promising tool for characterization of protein adsorption on arbitrary surfaces, but also the use of linear and nonlinear approaches in stationary phase characterization that strengthened drawn conclusions.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2015. 50 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 1248
National Category
Analytical Chemistry
Research subject
Chemistry with specialization in Analytical Chemistry
Identifiers
urn:nbn:se:uu:diva-249003 (URN)978-91-554-9235-9 (ISBN)
Public defence
2015-05-22, A1:111a, Husargatan 3, BMC, Uppsala, 13:00 (English)
Opponent
Supervisors
Available from: 2015-04-27 Created: 2015-04-09 Last updated: 2015-07-07

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Undin, TorgnyDahlin, AndreasHörnaeus, KatarinaBergquist, JonasBergström Lind, Sara

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