Chi-Ying Lee1, Ping Gong2, Lara J. Gamble3, David W. Grainger2, and David G. Castner4. (1) Chemical Engineering, University of Washington, Box 351750, Seattle, WA 98195-1750, (2) Chemistry, Colorado State University, Ft. Collins, CO 80523, (3) Bioengineering, University of Washington, William H. Foege Building, Box 355061, Seattle, WA 98195-5061, (4) Bio and Chemical Engineering, University of Washington, Box 351750, Seattle, WA 98195-1750
Direct capture of DNA targets from complex biological media is an important objective of DNA microarray and biosensor technologies. This goal can be hindered by non-specific adsorption of non-target DNA and proteins. Minimizing nonspecific adsorption and optimizing capture of low concentrations of DNA via hybridization requires a non-fouling background and optimization of coverage and orientation of DNA probes. We evaluated two diluent thiols [11-mercapto-1-undecanol (MCU) and 11-mercapto-undecyl tetra ethylene glycol (OEG)] on surfaces prepared using singled-stranded DNA containing a thiol anchor group (HS-ssDNA). These mixed DNA/alkylthiol monolayers were studied with X-ray photoelectron spectroscopy (XPS), near-edge X-ray absorption fine structure spectroscopy (NEXAFS), 32P-radiolabeling, and surface plasmon resonance (SPR). XPS and radiolabeling results show HS-ssDNA surface coverage decreases with extended exposure to either diluent. NEXAFS indicates increased order/upright orientation of the HS-ssDNA after 30 min of backfill. SPR was used to compare hybridization responses in various concentrations of serum. Although probe DNA surfaces with MCU and OEG diluents showed similar hybridization efficiency from target DNA in buffer, OEG incorporation into the DNA adlayer improved surface resistance to protein, allowing the detection of small DNA target sequences from undiluted, unpurified complex biological mixtures that was unachievable with DNA/MCU probe surfaces.