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Surface analysis of pure and complex mucin coatings
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
2009 (English)In: Journal of Colloid and Interface Science, ISSN 0021-9797, E-ISSN 1095-7103, Vol. 333, no 1, 180-187 p.Article in journal (Refereed) Published
Abstract [en]

In the past, we introduced the idea of using mucin coatings to improve biomaterials performance. Here, we evaluate non-radioactive methods for the analysis of pure and human host protein-containing (complex) mucin coatings on a real-type substrate (Thermanox). A common protein quantification assay (mBCA) was combined with mass-calibrated, enzyme-amplified assays based on lectin (ELLA) and antibody (ELISA)   recognition, to determine the total and specific amounts of surface-associated proteins. Model studies showed the mBCA assay to be of limited use at low mass loads, and steric effects to influence the ELLA at high surface layer densities. Non-specific responses due to   substrate interaction were low for the ELLA and ELISAs. Cross-reactions were observed during ELLA analysis of analytes sharing high degree of O-glycosylation. Combined mBCA-ELLA-ELISA analysis suggested that mucin desorption was low upon protein addition and that low concentrations of ELISA-determined Protein for the complex coatings Could be explained in terms of low accessibility of proteins to the bulk environment. Specifically, a methodology is presented for the determination of the fraction of surface-exposed, presumed bioactive proteins in a complex mucin coating. Finally, X-ray photoelectron spectroscopy and infrared reflectance spectroscopy combined with multivariate data analysis were proven useful in the evaluation of mucin-based coatings.

Place, publisher, year, edition, pages
2009. Vol. 333, no 1, 180-187 p.
Keyword [en]
Mucin quantification, mBCA, ELLA, ELISA, Ellipsometry, Jacalin, Complex mucin coating, Surface-exposed protein, Bioactivity, XPS, IRS-MVDA, Principal component analysis (PCA)
National Category
Chemical Sciences
Identifiers
URN: urn:nbn:se:uu:diva-97900DOI: 10.1016/j.jcis.2009.01.069ISI: 000264897200024OAI: oai:DiVA.org:uu-97900DiVA: diva2:173005
Available from: 2008-11-28 Created: 2008-11-28 Last updated: 2010-08-03Bibliographically approved
In thesis
1. On the Development of Mucin-based Biomaterial Coatings
Open this publication in new window or tab >>On the Development of Mucin-based Biomaterial Coatings
2008 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Owing to their key role in mucosal functioning as surface barriers with biospecific interaction potentials, the mucins are interesting candidates for use as surface modifiers in biomaterials applications.

In this work, “mild” fractionation procedures were used to prepare mucins of bovine (BSM), porcine (PGM), and human (MG1) origin. Biophysicochemical analysis showed the prepared mucins to differ in size, charge, conformation, and composition. In turn, these factors were shown to govern mucin adsorption on hydrophilic and hydrophobic model surfaces.

To enable for detailed coating analysis, methods for the qualitative and quantitative analysis of mucin-based coatings were developed. Of particular interest, a method for the determination of the fraction of surface-exposed, presumed bioactive proteins in a complex mucin coating was described.

It was shown, using microscopy and activation assays, that mucin precoating effectively suppresses the neutrophil response towards a polymeric model biomaterial. Under optimal coating conditions, all mucins performed equally well, thus indicating them to be functionally similar. Coating analysis suggested that efficient mucin surface-shielding is critical for good mucin coating performance.

Following a study on the complexation of albumin with preadsorbed mucin, we investigated the effect of mucin precoating on the conformation and neutrophil-activating properties of adsorbed host proteins. We found that mucin precoating greatly reduces the strong immune-response normally caused by adsorbed proinflammatory proteins (IgG and sIgA). Detailed coating analysis revealed that the fraction of surface-exposed protein in the mucin-protein composite influences the neutrophil response. Unexpectedly low neutrophil activation for composites containing near-monolayer concentrations of exposed IgG, suggested IgG to act synergistically with mucin on the surface. Conformational analysis supported this by showing that a preadsorbed mucin layer could stabilize adsorbed IgG through complexation. Our findings link well to the complex in vivo situation and suggest that functional mucosal mimics can be created in situ for improved biomaterials performance.

Place, publisher, year, edition, pages
Uppsala: Universitetsbiblioteket, 2008. 83 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 583
Keyword
Mucin, Biomaterial, Surface-exposed protein, XPS, Neutrophil, Cell morphology, SEM, QCM-D, Viscoelasticity, Protein-stabilization, HNL, Coating, MG1, BSM, PGM, SEC-MALS-RI, Mucin quantification, Protein adsorption, Ellipsometry
National Category
Dentistry
Identifiers
urn:nbn:se:uu:diva-9439 (URN)978-91-554-7368-6 (ISBN)
Public defence
2008-12-19, B21, BMC, Husargatan 3, 75123, Uppsala, 13:15
Opponent
Supervisors
Available from: 2008-11-28 Created: 2008-11-28 Last updated: 2010-12-14Bibliographically approved

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