David Kiaei

Tight binding of albumin to glow discharge treated polymers

Tetrafluoroethylene (TFE) glow discharge-treated Dacron vascular grafts resist thrombus deposition, embolization and thrombotic occlusion. In addition, albumin adsorbed on TFE-treated surfaces resists elution by sodium dodecyl sulfate (SDS). Since the tight binding of albumin to TFE-treated surfaces may contribute to their thromboresistant character, we decided to examine the mechanism responsible for this tenacious adsorption. We have investigated albumin adsorption and retention (after SDS elution) on a number of untreated and glow discharge-treated surfaces. Fluorocarbon glow discharge-treated polymers retain a larger fraction of the adsorbed albumin than ethylene and hexamethyldisiloxane glow discharge-treated surfaces. Albumin retention by surfaces appears to be closely related to their surface free energy (in air). Low energy surfaces (in air), whether untreated or glow discharge-treated, retain a larger fraction of the albumin adsorbed than higher energy surfaces. The lowest energy surfaces should have the highest interfacial energies in water, with correspondingly high driving forces for adsorption of proteins. This can lead to the formation of multiple binding sites upon adsorption, permitting strong hydrophobic interactions, which leads to the observed strong binding.

Activity of horseradish peroxide adsorbed on radio frequency glow discharge-treated polymers.

Horseradish peroxidase (HRP) has been used as a model enzyme in this study of its physical adsorption and residual enzyme activity on radio frequency glow discharge (RFGD)-treated polymers. The specific enzymatic activity of HRP adsorbed on different surfaces was assumed to be an indication of the extent of its conformational alterations on the surfaces. The surfaces studied were poly (ethylene terephthalate) (PET), polytetrafluoroethylene (PTFE), and tetrachloroethylene and tetrafluoroethylene glow discharge-treated PET, abbreviated as TCE/PET and TFE/PET. All surfaces were characterized by electron spectroscopy for chemical analysis (ESCA) and liquid contact angles in air. HRP adsorbs more strongly onto the two discharge-treated surfaces than onto the untreated polymers, as evidenced by the lower amount of HRP eluted by sodium dodecyl sulfate (SDS) from the treated polymers. For example, seventy percent of the HRP adsorbed on TCE/PET or TFE/PET remains on the surface after overnight elution with a 1% solution of SDS. In contrast, untreated PET and PTFE each retains only c. 20% of the absorbed enzyme. The enzymatic activity of HRP adsorbed on the different surfaces was studied using hydrogen peroxide (H2O2) as the substrate. HRP adsorbed on the higher energy surfaces, PET and TCE/PET, retains significantly more activity than the HRP adsorbed on the lower energy surfaces, PTFE and TFE/PET which appear to destroy rapidly almost all of the activity of HRP after it adsorbs. HRP adsorbed on TCE/PET is relatively more stable over time than HRP adsorbed on PET or free HRP in solution. (For example, only 45% of the specific enzymatic activity of HRP adsorbed on TCE/PET was lost after 3 h of storage in phosphate buffer at 37 degrees C, while 70% of that adsorbed on PET was lost.) In summary, when HRP is adsorbed on TCE/PET, it is very tightly bound, and yet it maintains a significant fraction of its initial specific activity and also retains this activity for 3 h in phosphate buffer at 37 degrees C. Thus, tenacious physical adsorption of proteins such as enzymes on TCE glow discharge-treated surfaces may have potential as a new method of immobilization of such molecules, for uses in biosensors, diagnostics, bioseparations, cell culture and bioreactors.

Radio-frequency gas discharge (RFGD) fluorination of polymers: Protein and cell interactions at RFGD-fluorinated interfaces

Abstract Proteins naturally adsorb at foreign surfaces. Adsorbed proteins on biomaterial surfaces are important to the biocompatibility of medical implants and devices, as well as to uses in biosensors, diagnostics, separations and industrial bioprocesses. There is ample evidence that the composition, organization, and conformations of the proteins adsorbed at a foreign interface are sensitive to the composition, topography, and molecular mobility of the substrate surface. The protein layer can also change in all the above aspects with time. Thus, it is a dynamic, “living” coating whose character is a direct reflection of the character of the substrate surface. Therefore, it is not surprising that many researchers have modified biomaterial surface compositions in order to influence protein absorption and subsequent conformational changes. The gas discharge process is one of the more useful methods for modification of biomaterial surfaces. Surfaces may be ablated (or etched) in a process which removes material and creates a cleaner, but chemically-modified surface. Gas discharge may also be used to deposit a polymer-like coating of a new composition on the surface, such as silicone and fluorocarbon coatings on hydrocarbon polymer surfaces. Certain of these fluorocarbon discharge-deposited polymers have been shown to enhance retention of adsorbed proteins, presumably by increased hydrophobic interactions with the modified substrate. Furthermore, platelet adhesion on such surfaces may be increasingly reduced as the adsorbed protein is increasingly bound to the fluorinated surface. These unusual results and their biologic implications are discussed in this paper.

Adsorption, retention and biologic activity of proteins adsorbed on gas discharge treated surfaces

We have observed that proteins adsorb very tenaciously to polymer surfaces that have been treated with certain gases in a gas discharge. These gases include tetrafluoroethylene (TFE) and tetrachloroethylene (TCE). We have studied adsorption of plasma proteins, antibodies, antigens and enzymes. In this article we present results on the adsorption, retention and biologic activity of (a) albumin as a passivating surface, (b) malaria and schistosomiasis antibodies in an immunoassay and (c) horseradish peroxidase (HRP) as a model enzyme biosensor. In some cases, the protein adsorbed to the treated surface retains greater activity and/or greater stability when compared to the untreated control surface. We also discuss the possible mechanism(s) of these interesting and unusual effects.