AVS 57th International Symposium & Exhibition
    Biomaterial Interfaces Wednesday Sessions
       Session BI-WeA

Paper BI-WeA11
Analysis of Unspecific Protein Adsorption onto Polymer Materials using Radioactive Labeling, Atomic Force Microscopy and ELISA

Wednesday, October 20, 2010, 5:20 pm, Room Taos

Session: Proteins & Peptides on Surfaces
Presenter: M. Holmberg, Technical University of Denmark
Authors: M. Holmberg, Technical University of Denmark
X.L. Hou, Technical University of Denmark
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In this study radioactive labeling is used in combination with ELISA measurements and Atomic Force Microscopy (AFM) analysis to investigate aspects of unspecific protein adsorption onto polymer materials. The radioactive labeling is a setup in which different proteins are labeled with isotopes that emit gamma radiation with different energies. This makes it possible to detect several proteins simultaneously onto the same sample and thus to investigate competitive protein adsorption and how the presence of some proteins influence the adsorption of others. Results from protein adsorption onto polymer materials using the radioactive labeling setup have shown adsorption levels higher than expected for monolayer adsorption and suggest the existence of protein multilayers on some surfaces. Results from fibrinogen adsorption onto surfaces that are pre-adsorbed with albumin show that fibrinogen can adsorb on top of albumin and that exchange of already adsorbed albumin is not a dominant process during the competitive adsorption with fibrinogen. Preliminary results on QCM-D (quartz crystal microbalance with dissipation monitoring) also strengthen the idea of the existence of an interface between polymer surface and protein solution in which proteins interact with both each other and the surface in a matrix structure that have multilayer character. To further illustrate the impact on (or lack of) unspecific protein adsorption using blocking buffers and pre-adsorption of proteins, we show results from ELISA measurements of unspecific protein adsorption onto TCPS (tissue culture polystyrene) and PS (polystyrene). We can detect large difference in adsorption level between proteins with and without a HIS tag (six histidines) onto TCPS, but not onto PS, with the HIS tagged proteins showing much higher adsorption onto TCPS compared to the same protein without a HIS tag. Furthermore, low or none impact on the level of adsorption of these HIS tagged proteins is observed when the TCPS surfaces are blocked with BSA (bovine serum albumin). We are combining quantitative results from radioactive labeling (and QCM-D) with AFM analysis performed in liquid to obtain data regarding homogeneity and topography of adsorbed protein layers. Furthermore, ELISA is used as a supplementary technique to acquire more knowledge regarding unspecific adsorption of proteins onto polymer materials. The obtained information is of importance when evaluating interactions between proteins and biomaterials.