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Interaction of human plasma proteins with thin gelatin-based hydrogel films: a QCM-D and ToF-SIMS study.

Schönwälder SM, Bally F, Heinke L, Azucena C, Bulut ÖD, Heißler S, Kirschhöfer F, Gebauer TP, Neffe AT, Lendlein A, Brenner-Weiß G, Lahann J, Welle A, Overhage J, Wöll C - Biomacromolecules (2014)

Bottom Line: This technique enables the determination of adsorbant mass and changes in the shear modulus of the hydrogel layer upon adsorption of human proteins.Furthermore, Secondary Ion Mass Spectrometry and principal component analysis was applied to monitor the changed composition of the topmost adsorbate layer.This approach opens interesting perspectives for a sensitive screening of viscoelastic biomaterials that could be used for regenerative medicine.

View Article: PubMed Central - PubMed

Affiliation: Karlsruhe Institute of Technology (KIT), Institute of Functional Interfaces (IFG) , 76344 Eggenstein-Leopoldshafen, Germany.

ABSTRACT
In the fields of surgery and regenerative medicine, it is crucial to understand the interactions of proteins with the biomaterials used as implants. Protein adsorption directly influences cell-material interactions in vivo and, as a result, regulates, for example, cell adhesion on the surface of the implant. Therefore, the development of suitable analytical techniques together with well-defined model systems allowing for the detection, characterization, and quantification of protein adsorbates is essential. In this study, a protocol for the deposition of highly stable, thin gelatin-based films on various substrates has been developed. The hydrogel films were characterized morphologically and chemically. Due to the obtained low thickness of the hydrogel layer, this setup allowed for a quantitative study on the interaction of human proteins (albumin and fibrinogen) with the hydrogel by Quartz Crystal Microbalance with Dissipation Monitoring (QCM-D). This technique enables the determination of adsorbant mass and changes in the shear modulus of the hydrogel layer upon adsorption of human proteins. Furthermore, Secondary Ion Mass Spectrometry and principal component analysis was applied to monitor the changed composition of the topmost adsorbate layer. This approach opens interesting perspectives for a sensitive screening of viscoelastic biomaterials that could be used for regenerative medicine.

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Adsorbed wet mass of Fbn on bare gold (dark gray), CVD (light gray),and gelatin film (white) calculated with the Voigt model from QCM-Dmeasurements performed at 37 °C.
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fig5: Adsorbed wet mass of Fbn on bare gold (dark gray), CVD (light gray),and gelatin film (white) calculated with the Voigt model from QCM-Dmeasurements performed at 37 °C.

Mentions: For quantitative protein/hydrogel interactionstudies using the QCM-D technology, very thin gelatin films (<100nm, swollen state) are mandatory to avoid strong damping of the shearwave oscillation by viscoelastic losses within the hydrogel layer.These requirements are met by the gelatin films produced via the reportedprotocol. Therefore, the protein/hydrogel interaction can be investigatedby QCM-D, where the resonance frequencies (Δf) and the energy dissipations (ΔD) are recordedduring the protein adsorption process. The resonance frequencies decreasewith increasing mass of adsorbed protein (m) perarea (A). The wet protein mass on the sensor crystalwas calculated from frequency and dissipation data applying the Voigtmodel.45,50 The frequency and dissipation shift duringFbn adsorption on three different coatings (gelatin, gold, CVD polymer)at the third overtone is shown in Figure 4.The corresponding calculated wet masses of the adsorbed proteins (HSAor Fbn) determined by the Voigt model are displayed in Figure 5. The adsorption of human protein at the surfaceof the gelatin-based hydrogel (7.9 ± 0.8 mg/m2, n = 13, for Fbn, and 0.4 ± 0.1 mg/m2, n = 8, for HSA) was much lower than at the CVD-modifiedsurface and gold (35.8 ± 0.9 mg/m2, n = 7, and 18.1 ± 1.0 mg/m2, n =5, respectively, for Fbn, and 11.4 ± 1.3 mg/m2, n = 12 and 12 ± 1.6 mg/m2, n = 5, respectively, for HSA). All experiments were repeated severaltimes. The standard deviations of the results of the individual experimentsare indicated by the error bars in Figure 5. It has to be noted that the statistical deviations are very lowand that the experiments are well reproducible. Compared to reportedresults on fibrinogen adsorbed on gold under similar conditions,51 our obtained values for ΔD and Δf lie in the expected range. Actuallythe absorbed amounts of Fbn and HSA were close to the amounts reportedfor PEG-modified surfaces (0.5–1.0 mg/m2 for HSAand 1.0–2.0 mg/m2 for Fbn)52 but still much higher than optimized PEG-based protein resistantsurfaces (<0.01 mg/m2 for HSA and ∼0.01–0.30mg/m2 for Fbn).26,53


Interaction of human plasma proteins with thin gelatin-based hydrogel films: a QCM-D and ToF-SIMS study.

Schönwälder SM, Bally F, Heinke L, Azucena C, Bulut ÖD, Heißler S, Kirschhöfer F, Gebauer TP, Neffe AT, Lendlein A, Brenner-Weiß G, Lahann J, Welle A, Overhage J, Wöll C - Biomacromolecules (2014)

Adsorbed wet mass of Fbn on bare gold (dark gray), CVD (light gray),and gelatin film (white) calculated with the Voigt model from QCM-Dmeasurements performed at 37 °C.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4215905&req=5

fig5: Adsorbed wet mass of Fbn on bare gold (dark gray), CVD (light gray),and gelatin film (white) calculated with the Voigt model from QCM-Dmeasurements performed at 37 °C.
Mentions: For quantitative protein/hydrogel interactionstudies using the QCM-D technology, very thin gelatin films (<100nm, swollen state) are mandatory to avoid strong damping of the shearwave oscillation by viscoelastic losses within the hydrogel layer.These requirements are met by the gelatin films produced via the reportedprotocol. Therefore, the protein/hydrogel interaction can be investigatedby QCM-D, where the resonance frequencies (Δf) and the energy dissipations (ΔD) are recordedduring the protein adsorption process. The resonance frequencies decreasewith increasing mass of adsorbed protein (m) perarea (A). The wet protein mass on the sensor crystalwas calculated from frequency and dissipation data applying the Voigtmodel.45,50 The frequency and dissipation shift duringFbn adsorption on three different coatings (gelatin, gold, CVD polymer)at the third overtone is shown in Figure 4.The corresponding calculated wet masses of the adsorbed proteins (HSAor Fbn) determined by the Voigt model are displayed in Figure 5. The adsorption of human protein at the surfaceof the gelatin-based hydrogel (7.9 ± 0.8 mg/m2, n = 13, for Fbn, and 0.4 ± 0.1 mg/m2, n = 8, for HSA) was much lower than at the CVD-modifiedsurface and gold (35.8 ± 0.9 mg/m2, n = 7, and 18.1 ± 1.0 mg/m2, n =5, respectively, for Fbn, and 11.4 ± 1.3 mg/m2, n = 12 and 12 ± 1.6 mg/m2, n = 5, respectively, for HSA). All experiments were repeated severaltimes. The standard deviations of the results of the individual experimentsare indicated by the error bars in Figure 5. It has to be noted that the statistical deviations are very lowand that the experiments are well reproducible. Compared to reportedresults on fibrinogen adsorbed on gold under similar conditions,51 our obtained values for ΔD and Δf lie in the expected range. Actuallythe absorbed amounts of Fbn and HSA were close to the amounts reportedfor PEG-modified surfaces (0.5–1.0 mg/m2 for HSAand 1.0–2.0 mg/m2 for Fbn)52 but still much higher than optimized PEG-based protein resistantsurfaces (<0.01 mg/m2 for HSA and ∼0.01–0.30mg/m2 for Fbn).26,53

Bottom Line: This technique enables the determination of adsorbant mass and changes in the shear modulus of the hydrogel layer upon adsorption of human proteins.Furthermore, Secondary Ion Mass Spectrometry and principal component analysis was applied to monitor the changed composition of the topmost adsorbate layer.This approach opens interesting perspectives for a sensitive screening of viscoelastic biomaterials that could be used for regenerative medicine.

View Article: PubMed Central - PubMed

Affiliation: Karlsruhe Institute of Technology (KIT), Institute of Functional Interfaces (IFG) , 76344 Eggenstein-Leopoldshafen, Germany.

ABSTRACT
In the fields of surgery and regenerative medicine, it is crucial to understand the interactions of proteins with the biomaterials used as implants. Protein adsorption directly influences cell-material interactions in vivo and, as a result, regulates, for example, cell adhesion on the surface of the implant. Therefore, the development of suitable analytical techniques together with well-defined model systems allowing for the detection, characterization, and quantification of protein adsorbates is essential. In this study, a protocol for the deposition of highly stable, thin gelatin-based films on various substrates has been developed. The hydrogel films were characterized morphologically and chemically. Due to the obtained low thickness of the hydrogel layer, this setup allowed for a quantitative study on the interaction of human proteins (albumin and fibrinogen) with the hydrogel by Quartz Crystal Microbalance with Dissipation Monitoring (QCM-D). This technique enables the determination of adsorbant mass and changes in the shear modulus of the hydrogel layer upon adsorption of human proteins. Furthermore, Secondary Ion Mass Spectrometry and principal component analysis was applied to monitor the changed composition of the topmost adsorbate layer. This approach opens interesting perspectives for a sensitive screening of viscoelastic biomaterials that could be used for regenerative medicine.

Show MeSH
Related in: MedlinePlus