Methods to manipulate and visualize isolated DNA and oligonucleotide strands are important for investigation of their biophysics as well as their interactions with proteins. Herein, we report such a method by combining a block copolymer surface functionalization strategy with microfluidics. The copolymer poly(L-lysine-graftpolyethylene glycol) (PLL-g-PEG) coated one surface of the microfluidic channels, rendering it passive to adsorption and thus minimizing any noise arising from nontargeted adsorbed molecules. Single λ-phage DNA molecules were immobilized and were extended by molecular combing. Their extension did not exceed their contour length, which we attribute to the low surface tension of the coated surface. To demonstrate further the applicability of our method, the anchored DNA was extended by hydrodynamic flow. We propose this method for exploring DNA-protein interactions due to the copolymer's enhanced capacity for single-molecule detection, stability under wet or dry conditions, hydrophilicity, full compatibility with microfluidics and simplicity being a one-step process. © 2010 American Chemical Society.