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Undin, T., Dahlin, A. P., Bergquist, J. & Bergström Lind, S. (2018). Mass Spectrometric Determination of the Effect of Surface Deactivation on Membranes Used for In-Situ Sampling of Cerebrospinal Fluid (CSF). Separations, 5(2), Article ID 27.
Open this publication in new window or tab >>Mass Spectrometric Determination of the Effect of Surface Deactivation on Membranes Used for In-Situ Sampling of Cerebrospinal Fluid (CSF)
2018 (English)In: Separations, ISSN 2297-8739, Vol. 5, no 2, article id 27Article in journal (Refereed) Published
Abstract [en]

In this paper, a strategy for structured monitoring of surface modifications to control protein adsorption to membrane structures is presented. The already established on-surface enzymatic digestion (oSED) method combined with nano-liquid chromatography and tandem mass spectrometry (LC-MS/MS) analysis was employed for the analysis of proteins in ventricular cerebrospinal fluid (vCSF) from neurointensive care patients. Protein adsorption was studied by in-situ sampling in a temporally resolved manner on both immobilized native and Pluronic-deactivated membranes. Deactivation was significantly reducing the protein adsorption but it also induced novel selective properties of the surface. The proposed versatile strategy will facilitate protein-biomaterial, protein-polymer, protein-protein interaction studies in the future.

Keywords
Protein Adsorption, Mass Spectrometry, oSED, Surface modification, Coating, Poloxamer 407, Pluronic F127, Trypsin, Digestion, Shotgun, Bottom-up, peptides
National Category
Analytical Chemistry Engineering and Technology
Research subject
Chemistry with specialization in Analytical Chemistry; Chemistry with specialization in Surface Biotechnology
Identifiers
urn:nbn:se:uu:diva-248631 (URN)10.3390/separations5020027 (DOI)000436515600007 ()
Funder
Magnus Bergvall Foundation, 2015-01200Åke Wiberg Foundation, M14-0127Swedish Research Council, 2015-4870Magnus Bergvall Foundation, 2016-01675Carl Tryggers foundation , CST 15:57Berzelii Centre EXSELENT
Available from: 2015-04-05 Created: 2015-04-05 Last updated: 2018-10-12Bibliographically approved
Undin, T., Dahlin, A., Hörnaeus, K., Bergquist, J. & Bergström Lind, S. (2016). Mechanistic investigation of the on surface enzymatic digestion (oSED) protein adsorption detection method using targeted mass spectrometry. The Analyst, 141(5), 1714-1720
Open this publication in new window or tab >>Mechanistic investigation of the on surface enzymatic digestion (oSED) protein adsorption detection method using targeted mass spectrometry
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2016 (English)In: The Analyst, ISSN 0003-2654, E-ISSN 1364-5528, Vol. 141, no 5, p. 1714-1720Article 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.

National Category
Analytical Chemistry Engineering and Technology
Research subject
Chemistry with specialization in Analytical Chemistry
Identifiers
urn:nbn:se:uu:diva-248629 (URN)10.1039/c5an02091c (DOI)000371229600018 ()
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
Chu, J., Undin, T., Lind, S., Dahlin, A. & Hjort, K. (2015). Influence of different pluronic surface modifications and pressure on microdialysis protein extraction efficiency. Biomedical microdevices (Print)
Open this publication in new window or tab >>Influence of different pluronic surface modifications and pressure on microdialysis protein extraction efficiency
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2015 (English)In: Biomedical microdevices (Print), ISSN 1387-2176, E-ISSN 1572-8781Article in journal (Refereed) Submitted
Abstract [en]

There is growing interest in using microdialysis (MD) for monitoring larger and more complexmolecules such as neuropeptides and proteins. This promotes the use of MD membranes withmolecular weight cut off (MWCO) of 100 kDa. Hence, the hydrodynamic property of themembrane goes to ultrafiltration, making the sampling more sensitive to pressure changes. Also,despite the large membrane pore size, studies have shown that membrane biofouling still leads tounstable catheter performance. Our objective is to study in vitro how four kinds of surfacemodifications (Pluronic L31, L44, F87 and F127+L31) affect the fluid recovery (FR) andextraction efficiency (EE) of 100 kDa MWCO MD catheters, under controlled pressure. Apressure chamber was employed to facilitate the tests, using as MD sample a protein standardwith proteins of similar concentrations as in human cerebral spinal fluid. The collected dialysatefractions were examined for FR and EE. Targeted mass spectrometry analysed the EE ofindividual proteins and peptides. The thicker the pluronic adsorption layer, the less thehydrodynamic diameter of the membrane pores, leading to lower and more stable FR. The foursurface modifications had three different behaviours: Pluronic F127 + L31 showed similarbehavior to the Pluronic F127 and the native original membrane; Pluronic F87 showed acontinuous EE increase with pressure; Pluronic L31 and L44 showed similar EE values, whichwere stable with pressure. Different surface modifications are clearly selective to differentproteins and peptides. We conclude that a pluronic surface modification could provide MDsampling with more stable FR, and more stable or enhanced EE with high FR, depending on theobjective of the sampling.

Place, publisher, year, edition, pages
Springer: , 2015
Keywords
microdialysis, surface modification, poloxamer, protein, extraction efficiency
National Category
Nano Technology
Research subject
Engineering Science with specialization in Microsystems Technology; Engineering Science with specialization in Materials Science
Identifiers
urn:nbn:se:uu:diva-261557 (URN)
Funder
Berzelii Centre EXSELENT
Available from: 2015-09-01 Created: 2015-09-01 Last updated: 2017-12-04Bibliographically approved
Chu, J., Undin, T., Bergström Lind, S., Hjort, K. & Dahlin, A. (2015). Influence of surface modification and static pressure on microdialysis protein extraction efficiency. Biomedical microdevices (Print), 17(5), Article ID UNSP 96.
Open this publication in new window or tab >>Influence of surface modification and static pressure on microdialysis protein extraction efficiency
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2015 (English)In: Biomedical microdevices (Print), ISSN 1387-2176, E-ISSN 1572-8781, Vol. 17, no 5, article id UNSP 96Article in journal (Refereed) Published
Abstract [en]

There is growing interest in using microdialysis (MD) for monitoring larger and more complexmolecules such as neuropeptides and proteins. This promotes the use of MD membranes withmolecular weight cut off (MWCO) of 100 kDa or above. The hydrodynamic property of themembrane goes to ultrafiltration or beyond, making the MD catheters more sensitive to pressure.In the meantime, despite the large pore size, studies have shown that membrane biofouling stilllead to unstable catheter performance. The objective is to study in vitro how 500 kDa dextranand Poloxamer 407 surface modification affect the fluid recovery (FR) and extraction efficiency(EE) of 100 kDa MWCO MD catheters. A pressure chamber was designed to facilitate the tests,using as MD sample a protein standard with similar concentrations as in human cerebral spinalfluid, comparing native and Poloxamer 407 modified MD catheters. The collected dialysatefractions were examined for FR and protein EE, employing Dot-it Spot-it Protein Assay for totalprotein EE and targeted mass spectrometry (MS) for EE of individual proteins and peptides. TheFR results suggested that the surface modified catheters were less sensitive to the pressure andprovide higher precision, and provided a FR closer to 100%. The surface modification did notshow a significant effect on the protein EE. The average total protein EE of surface modifiedcatheters was slightly higher than that of the native ones. The MS EE data of individual proteinsshowed a clear trend of complex response in EE with pressure.

Place, publisher, year, edition, pages
Springer: , 2015
Keywords
microdialysis, surface modification, poloxamer, protein, extraction efficiency
National Category
Nano Technology
Research subject
Engineering Science with specialization in Microsystems Technology
Identifiers
urn:nbn:se:uu:diva-261031 (URN)10.1007/s10544-015-0005-3 (DOI)000362281200013 ()26342494 (PubMedID)
Funder
Berzelii Centre EXSELENTSwedish Research Council, P29797-1Åke Wiberg FoundationMagnus Bergvall FoundationScience for Life Laboratory - a national resource center for high-throughput molecular bioscience
Available from: 2015-08-28 Created: 2015-08-28 Last updated: 2017-12-04Bibliographically approved
Undin, T., Bergström Lind, S. & Dahlin, A. P. (2015). MS for investigation of time-dependent protein adsorption on surfaces in complex biological samples. Future Science OA, Article ID 32.
Open this publication in new window or tab >>MS for investigation of time-dependent protein adsorption on surfaces in complex biological samples
2015 (English)In: Future Science OA, ISSN 2056-5623, article id 32Article in journal (Refereed) Published
Abstract [en]

Aim: This study aims at developing a nondestructive way for investigating proteinadsorption on surfaces such as biomaterials using mass spectrometry. Methods: Ventricular cerebrospinal fluid in contact with poly carbonate membranes were usedas adsorption templates and on-surface enzymatic digestion was applied to desorbproteins and cleave them into peptides. Mass spectrometric analysis provided bothprotein identification and determination of protein specific adsorption behavior. Results: In general, the adsorption increased with incubation time but also proteinspecifictime-resolved adsorption patterns from the complex protein solutionwere discovered. Conclusion: The method developed is a promising tool for thecharacterization of biofouling, which sometimes causes rejection and encapsulationof implants and can be used as complement to other surface analytical techniques.

One problem associated with artificial materials in the body is that proteins in thebody interact with the surface, which sometimes causes rejection of the implant.In this study, a method for investigating the time-dependent protein adsorptionon a surface originating from a complex biological protein solution was developed.Compared with other surface analyses, this method can identify what proteins thatadsorbs on the surface. In addition, determination of protein-specific adsorptionbehavior in relation to incubation was possible

National Category
Chemical Sciences Engineering and Technology
Identifiers
urn:nbn:se:uu:diva-262022 (URN)10.4155/fso.15.32 (DOI)
Funder
Åke Wiberg Foundation, M14–0127Magnus Bergvall Foundation, 2014-00367
Available from: 2015-09-07 Created: 2015-09-07 Last updated: 2017-01-24Bibliographically approved
Chu, J., Undin, T., Dahlin, A., Wang, C., Park, J. & Hjort, K. (2015). Protein Desalination Chip for Mass Spectrometry Sample Preparation. In: : . Paper presented at The 19th International Conference on Miniaturized Systems for Chemistry and Life Sciences, MicroTAS 2015. October 25-29, 2015. Gyeongju, Korea.. MicroTAS
Open this publication in new window or tab >>Protein Desalination Chip for Mass Spectrometry Sample Preparation
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2015 (English)Conference paper, Oral presentation with published abstract (Refereed)
Abstract [en]

This work focuses on desalination of a protein sample in a lab-on-chip device using the ion concentration polarization (ICP) technique. It was demonstrated with a salt containing buffer with four proteins and two peptides of concentrations typical to cerebrospinal fluid (CSF). Not only was the output desalinated but its protein concentration with large molecular weight (MW) was as much as 3 times higher for the largest protein compared to the original. We conclude that ICP based microfluidic chips have great potential for desalination and protein concentration in microdialysis sampling coupled to mass spectroscopy (MS).

Place, publisher, year, edition, pages
MicroTAS: , 2015
Keywords
Ion concentration polarization, Desalination, Protein, Mass spectroscopy
National Category
Nano Technology
Research subject
Engineering Science with specialization in Microsystems Technology; Engineering Science with specialization in Materials Science
Identifiers
urn:nbn:se:uu:diva-261067 (URN)
Conference
The 19th International Conference on Miniaturized Systems for Chemistry and Life Sciences, MicroTAS 2015. October 25-29, 2015. Gyeongju, Korea.
Available from: 2015-08-28 Created: 2015-08-28 Last updated: 2016-01-05
Hillered, L., Dahlin, A. P., Clausen, F., Chu, J., Bergquist, J., Hjort, K., . . . Lewén, A. (2014). Cerebral microdialysis for protein biomarker monitoring in the neurointensive care setting - a technical approach. Frontiers in Neurology, 5, 245
Open this publication in new window or tab >>Cerebral microdialysis for protein biomarker monitoring in the neurointensive care setting - a technical approach
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2014 (English)In: Frontiers in Neurology, ISSN 1664-2295, E-ISSN 1664-2295, Vol. 5, p. 245-Article in journal (Refereed) Published
Abstract [en]

Cerebral microdialysis (MD) was introduced as a neurochemical monitoring method in the early 1990s and is currently widely used for the sampling of low molecular weight molecules, signaling energy crisis, and cellular distress in the neurointensive care (NIC) setting. There is a growing interest in MD for harvesting of intracerebral protein biomarkers of secondary injury mechanisms in acute traumatic and neurovascular brain injury in the NIC community. The initial enthusiasm over the opportunity to sample protein biomarkers with high molecular weight cut-off MD catheters has dampened somewhat with the emerging realization of inherent methodological problems including protein-protein interaction, protein adhesion, and biofouling, causing an unstable in vivo performance (i.e., fluid recovery and extraction efficiency) of the MD catheter. This review will focus on the results of a multidisciplinary collaborative effort, within the Uppsala Berzelii Centre for Neurodiagnostics during the past several years, to study the features of the complex process of high molecular weight cut-off MD for protein biomarkers. This research has led to new methodology showing robust in vivo performance with optimized fluid recovery and improved extraction efficiency, allowing for more accurate biomarker monitoring. In combination with evolving analytical methodology allowing for multiplex biomarker analysis in ultra-small MD samples, a new opportunity opens up for high-resolution temporal mapping of secondary injury cascades, such as neuroinflammation and other cell injury reactions directly in the injured human brain. Such data may provide an important basis for improved characterization of complex injuries, e.g., traumatic and neurovascular brain injury, and help in defining targets and treatment windows for neuroprotective drug development.

National Category
Neurology Engineering and Technology
Identifiers
urn:nbn:se:uu:diva-239227 (URN)10.3389/fneur.2014.00245 (DOI)25520696 (PubMedID)
Available from: 2014-12-19 Created: 2014-12-19 Last updated: 2017-12-05Bibliographically approved
Chu, J., Koudriavtsev, V., Hjort, K. & Dahlin, A. P. (2014). Flourescence imaging of molecule transport in high molecular weight cut-off microdialysis. In: : . Paper presented at 10th Micronano Systems Workshop (MSW 2014, 15-16 May, Uppsala, Sweden) (pp. 249-252).
Open this publication in new window or tab >>Flourescence imaging of molecule transport in high molecular weight cut-off microdialysis
2014 (English)Conference paper, Poster (with or without abstract) (Refereed)
National Category
Other Medical Engineering
Identifiers
urn:nbn:se:uu:diva-239237 (URN)
Conference
10th Micronano Systems Workshop (MSW 2014, 15-16 May, Uppsala, Sweden)
Available from: 2014-12-19 Created: 2014-12-19 Last updated: 2016-04-22
Chu, J., Koudriavtsev, V., Hjort, K. & Dahlin, A. P. (2014). Fluorescence imaging of macromolecule transport in high molecular weight cut-off microdialysis. Analytical and Bioanalytical Chemistry, 406(29), 7601-7609
Open this publication in new window or tab >>Fluorescence imaging of macromolecule transport in high molecular weight cut-off microdialysis
2014 (English)In: Analytical and Bioanalytical Chemistry, ISSN 1618-2642, E-ISSN 1618-2650, Vol. 406, no 29, p. 7601-7609Article in journal (Refereed) Published
Abstract [en]

When microdialysis (MD) membrane exceeds molecular weight cut-off (MWCO) of 100 kDa, the fluid mechanics are in the ultrafiltration regime. Consequently, fluidic mass transport of macromolecules in the perfusate over the membrane may reduce the biological relevance of the sampling and cause an inflammatory response in the test subject. Therefore, a method to investigate the molecular transport of high MWCO MD is presented. An in vitro test chamber was fabricated to facilitate the fluorescent imaging of the MD sampling process, using fluoresceinylisothiocyanate (FITC) dextran and fluorescence microscopy. Qualitative studies on dextran behavior inside and outside the membrane were performed. Semiquantitative results showed clear dextran leakage from both 40 and 250 kDa dextran when 100 kDa MWCO membranes were used. Dextran 40 kDa leaked out with an order of magnitude higher concentration and the leakage pattern resembled more of a convective flow pattern compared with dextran 250 kDa, where the leakage pattern was more diffusion based. No leakage was observed when dextran 500 kDa was used as a colloid osmotic agent. The results in this study suggest that fluorescence imaging could be used as a method for qualitative and semiquantitative molecular transport and fluid dynamics studies of MD membranes and other hollow fiber catheter membranes.

National Category
Other Medical Engineering Engineering and Technology
Identifiers
urn:nbn:se:uu:diva-239233 (URN)10.1007/s00216-014-8192-y (DOI)000344860300019 ()25286875 (PubMedID)
Available from: 2014-12-19 Created: 2014-12-19 Last updated: 2017-12-05Bibliographically approved
Wetterhall, M., Bergquist, J., Hillered, L., Hjort, K. & Dahlin, A. P. (2014). Identification of human cerebrospinal fluid proteins and their distribution in an in vitro microdialysis sampling system. European Journal of Pharmaceutical Sciences, 57(SI), 34-40
Open this publication in new window or tab >>Identification of human cerebrospinal fluid proteins and their distribution in an in vitro microdialysis sampling system
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2014 (English)In: European Journal of Pharmaceutical Sciences, ISSN 0928-0987, E-ISSN 1879-0720, Vol. 57, no SI, p. 34-40Article in journal (Refereed) Published
Abstract [en]

A qualitative study is presented on how proteins from a complex biological sample are distributed in a microdialysis sample system. A comparison between proteins identified in the human ventricular cerebrospinal fluid (CSF) sample, the collected dialysate and the proteins adsorbed onto the membrane was conducted. The microdialysis experiment was performed in vitro at 37 °C for the duration of 24h. Thereafter, the membranes were removed from the catheter and the adsorbed proteins were tryptically digested using the on-surface enzymatic digestion (oSED) protocol. The CSF samples and the dialysates were digested using a standard in-solution trypsin digestion protocol.  In the final phase, the samples were analysed using nano-liquid chromatography in combination with tandem mass spectrometry. In the four sample compartments analysed (CSF start, Membrane, Dialysate, CSF end) a total of 134 different proteins were found. However, most of the identified proteins(n=87) were uniquely found in one sample compartment only. Common CSF proteins such as albumin, apolipoproteins and cystatin C together with plasma proteins such as hemoglobin and fibrinogen were among the 11 proteins that were found in all samples. These proteins are present in high concentrations in CSF, which means that they effectively block out the detection signal of less abundant proteins.  Therefore, only 25 % of the proteins adsorbed onto the membrane were detected in the CSF compared with the dialysate that shared 44% of its proteins with the CSF. The proteins adsorbed onto the membrane were significantly more hydrophobic, had a lower instability index and more thermostable compared to the proteins in the CSF and the dialysate. The results suggest that proteins adsorbed onto the microdialysis membranes may escape detection because they are prevented from passing the membrane into the dialysate. Thus, the membrane needs to be examined after sample collection in order to better verify the protein content in the original sample. This is particularly important when searching for new protein biomarkers for neurodegenerative diseases.

National Category
Analytical Chemistry Engineering and Technology
Research subject
Engineering Science with specialization in Microsystems Technology
Identifiers
urn:nbn:se:uu:diva-212733 (URN)10.1016/j.ejps.2013.12.011 (DOI)000336471400005 ()
Available from: 2013-12-13 Created: 2013-12-13 Last updated: 2017-12-06Bibliographically approved
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ORCID iD: ORCID iD iconorcid.org/0000-0002-0041-0902

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