(77d) Protein Recognitive Hydrogel Systems for Biosensor Applications

Kryscio, D. R. - Presenter, University of Texas at Austin
Peppas, N. A. - Presenter, University of Texas at Austin

The objective of this research is to develop biomimetic recognitive hydrogel polymer networks that can be applied to biosensing or other nanoscale applications when integrated with a nanodevice. Molecular recognition or molecular imprinting is an emerging field of interest in which a polymer network is formed with specific recognition for a desired template molecule. Briefly, functional monomers are chosen which exhibit chemical structures designed to interact with the template molecule via covalent or non-covalent chemistry. The monomers are then polymerized in the presence of the desired template, the template is subsequently removed, and the product is a polymer with binding sites specific to the template molecule. This technique has been successfully applied to small molecule templates in the areas of separations, solid-phase extractions, artificial enzymes, and chemical sensors.

The ability to selectively recognize a specific protein in a complex solution (such as blood) would have many applications, including serving as a biosensor. Protein imprinted polymers are ideal replacements to their biological counterparts as they can be easily tailored to a variety of templates, are inexpensive and straightforward to prepare, have greater stability in harsh conditions, and are reusable. Because of this, protein imprinting has gained a great deal of attention from the scientific community; however, efforts to do so have achieved limited success due to the inherent properties of proteins, which include size, complexity, conformation, and solubility.

Protein imprinted polymers were synthesized via a thermal free radical polymerization of functional monomers methacrylic acid (MAA), acrylamide (Aam), and 2-(dimethylamino)ethyl methacrylate (DMAEMA) grafted with polyethylene glycol (400) dimethacrylate as the cross-linking monomer. Bovine serum albumin (BSA) was used as the model protein template. Electrostatic interactions were exploited between the charged functional monomers MAA and DMAEMA and the corresponding oppositely charged amino acids present in BSA. In addition, BSA contains polar, uncharged amino acids that can undergo hydrogen bonding with Aam. These components were dissolved in water to form the pre-polymerization solution, at which time the template and monomers were allowed to complex for 30 minutes in order for these interactions to occur. After purging with N2 to remove the free radical scavenger O2, Ammonium Persulfate (APS) and N,N,N',N'-tetramethylethylenediamine (TEMED) were added as the initiator and catalyst. The reaction proceeded overnight at room temperature. Control polymers were synthesized under exactly the same conditions, without the addition of BSA.

A mesh size of 48.1±9.0 nm for the molecularly imprinted polymer (MIP) and 44.6±11.5 nm for the non-imprinted polymer (NIP) was determined using the Peppas-Merrill equation. These values show that there is no discernable difference in mesh size with the imprinted and control samples, but more importantly they clearly demonstrate that the pore sizes of the hydrogel networks are large enough for easy diffusion of the 14 nm diameter BSA template molecule. Also, removal of BSA after polymerization has been monitored via UV/vis spectroscopy. We have demonstrated the ability to consistently remove BSA, 70-97%, at amounts near and above values in literature. Initial recognition studies have shown that the synthesized protein imprinted polymers have BSA recognition capabilities as 8.8 times more BSA was present in the elution phase of the MIP sample (13.8±4.5 mg) compared to that of the NIP (1.6±1.1mg).