Lithographic Patterning of Photoreactive Cell-Adhesive Proteins

We describe a novel, simple method for the photolithographic patterning of cell-adhesive proteins. Intrinsically photoreactive proteins are synthesized in Escherichia coli through incorporation of the non-canonical, photosensitive amino acid para-azidophenylalanine. Upon ultraviolet irradiation at 365 nm, proteins form cross-linked films with elastic moduli that can be tuned by varying the concentration of photoreactive amino acid in the expression medium. Films of these proteins can be directly patterned using standard photolithographic techniques. We demonstrate the utility of this method of protein patterning by creating stable arrays of fibroblast cells on an engineered protein “photoresist”

Physical properties of artificial extracellular matrix protein films prepared by isocyanate crosslinking

Artificial extracellular matrix proteins, genetically engineered from elastin- and fibronectin-derived repeating units, were crosslinked with hexamethylene diisocyanate in dimethylsulfoxide. The resulting hydrogel films were transparent, uniform, and highly extensible. Their tensile moduli depended on crosslinker concentration and spanned the range characteristic of native elastin. The water content of the films was low (∼27%), but the temperature-dependent swelling behavior of the crosslinked materials was reminiscent of the lower critical solution temperature property of the soluble polymers

Mechanically Tunable Thin Films of Photosensitive Artificial Proteins: Preparation and Characterization by Nanoindentation

Thin films of controlled elastic modulus were made by photo-cross-linking artificial extracellular matrix (aECM) proteins containing the photosensitive amino acid p-azidophenylalanine (pN_3Phe). The elastic moduli of the films were calculated from nanoindentation data collected by atomic force microscopy (AFM) using a thin-film Hertz model. The modulus was shown to be tunable in the range 0.3−1.0 MPa either by controlling the irradiation time or by varying the level of pN_3Phe in the protein. Tensile measurements on bulk films of the same proteins and finite-element simulation of the indentation process agreed with the thin-film modulus measurements from AFM. Substrates characterized by spatial variation in elastic modulus were created by local control of the irradiation time

Salicylic acid-releasing polyurethane acrylate polymers as anti-biofilm urological catheter coatings

Biofilm-associated infections are a major complication of implanted and indwelling medical devices like urological and venous catheters. They commonly persist even in the presence of an oral or intravenous antibiotic regimen, often resulting in chronic illness. We have developed a new approach to inhibiting biofilm growth on synthetic materials through controlled release of salicylic acid from a polymeric coating. Herein we report the synthesis and testing of a ultraviolet-cured polyurethane acrylate polymer composed, in part, of salicyl acrylate, which hydrolyzes upon exposure to aqueous conditions, releasing salicylic acid while leaving the polymer backbone intact. The salicylic acid release rate was tuned by adjusting the polymer composition. Anti-biofilm performance of the coatings was assessed under several biofilm forming conditions using a novel combination of the MBEC Assay™ biofilm multi-peg growth system and bioluminescence monitoring for live cell quantification. Films of the salicylic acid-releasing polymers were found to inhibit biofilm formation, as shown by bioluminescent and GFP reporter strains of Pseudomonas aeruginosaand Escherichia coli. Urinary catheters coated on their inner lumens with the salicylic acid-releasing polymer significantly reduced biofilm formation by E. coli for up to 5 days under conditions that simulated physiological urine flo