Glow-discharge radio frequency plasma polyether and fluoropolyether coated substrates prevent IgG protein adsorption

Abstract

Event Abstract Back to Event Glow-discharge radio frequency plasma polyether and fluoropolyether coated substrates prevent IgG protein adsorption Marvin M. Mecwan1* and Buddy Ratner1* 1 University of Washington, Bioengineering, United States Introduction: Protein-based pharmaceuticals present unique challenges in processing, packaging and delivery. All proteins rapidly adsorb to solid surfaces, essentially irreversible, and frequently leads to denaturation or aggregation of the protein. Also, there are concerns with substances from the packaging leaching out and affecting proteins. This study examines fundamental aspects of protein interactions with surfaces, particularly using glow discharge plasma-treated surfaces which can be readily applied to delivery devices, packaging and processing equipment [1],[2] and may lead to a new generation of surfaces effective for protein manufacture, storage and delivery. This research particularly focuses on plasma tetraglyme (TG), acrylic acid (AA), perfluoropropylene (C3F6), and perfluoromethyl vinyl ether (C3F6O) plasma treated glass, stainless steel, and cyclic olefin polymer (COP) substrates and their interaction with IgG. Methods: Glass and COP (8mm φ discs), and 316L stainless steel (7 x 7 mm2) substrates were sequentially cleaned with DCM, acetone, and methanol in a sonication bath for 10 mins x 2. RF-Plasma deposition and delamination: Substrates were loaded into the reactor, and argon etched (40W for 5 min). Using a mass flow controller, the monomer of choice was introduced into the chamber and were plasma deposited. Plasma-treated samples (n=3/group) were washed using DI water x 3 over 24 hours to assess whether coatings would delaminate. ESCA analyses were done using an S-Probe ESCA (with monochromatic Al K-alpha X-rays focused to 800µm spot size) using survey and detailed C1s scans. Data was analyzed using ESCA analysis software. IgG adsorption studies: Bovine IgG was tagged using an ICl method. Plasma-treated samples (n=3/treatment group) were immersed in 0.1mg/mL I-125 tagged bovine IgG solution in 1x cPBS with sodium iodide (cPBSI) for 1 hr. Samples were rinsed with cPBS x 3 and counted for 10 mins using a gamma counter. Cytotoxicity studies: Plasma-coated samples were eluted in complete growth medium for 24 hrs (370C, 5% CO2­ incubator). ~80% confluent NIH-3T3 fibroblasts were exposed to eluates from the plasma coated substrates, and observed over 48 hrs using an inverted microscope. ISO 10993-5 guidelines were used for cytotoxic evaluation. Results and Discussion: ESCA scans of plasma-treated substrates showed absence of substrate associated peaks (such as Fe or Si), suggesting that coatings did not delaminate; also implies that plasma coatings on substrates are at least 10 nm thick (ESCA resolution is 100Å). Furthermore, experimental and theoretical elemental compositions of the surfaces align well. Protein adsorption studies show that tetraglyme and acrylic acid plasma coatings are essentially non-fouling (<2 ng/cm2) as compared to C3F6 and C3F6O plasma coatings which adsorbed proteins in the range of 10-14 ng/cm2. Furthermore, all plasma treated substrates show reduced protein adsorption compared to untreated controls. Moreover, fibroblasts exposed to all plasma coating eluates remained viable after 48 hours. Conclusions: The data demonstrate that plasma-treated polyether and fluoropolyether substrates can create a new generation of surfaces that can be used for contact with protein therapeutic agents.Future studies will be aimed at investigating the strength of bonding of IgG to these surfaces, as well as protein aggregation studies. Winston Ciridon; Xia Dong, PhD