Fabrication of nanoporous thin-ﬁlm working electrodes and their biosensing applications Tingjie Li a , Falong Jia b,c , Yaxi Fan a , Zhifeng Ding b , Jun Yang a,n a Department of Mechanical and Materials Engineering, The University of Western Ontario, London, Ontario, Canada N6A 5B9 b Department of Chemistry, The University of Western Ontario, 1151 Richmond Street, London, Ontario, Canada N6A 5B7 c College of Chemistry, Central China Normal University, Wuhan 430079, PR China article info Article history: Received 16 July 2012 Received in revised form 19 September 2012 Accepted 2 October 2012 Available online 16 October 2012 Keywords: Nanoporous Electrochemical detection of glucose Biosensor Point-of-care abstract Electrochemical detection for point-of-care diagnostics is of great interest due to its high sensitivity, fast analysis time and ability to operate on a small scale. Herein, we report the fabrication of a nanoporous thin-ﬁlm electrode and its application in the conﬁguration of a simple and robust enzymatic biosensor. The nanoporous thin-ﬁlm was formed in a planar gold electrode through an alloying/dealloying process. The nanoporous electrode has an electroactive surface area up to 40 times higher than that of a ﬂat gold electrode of the same size. The nanoporous electrode was used as a substrate to build an enzymatic electrochemical biosensor for the detection of glucose in standard samples and control serum samples. The example glucose biosensor has a linear response up to 30 mM, with a high sensitivity of 0.50 mA mM 1 mm 2 , and excellent anti-interference ability against lactate, uric acid and ascorbic acid. Abundant catalyst and enzyme were stably entrapped in the nanoporous structure, leading to high stability and reproducibility of the biosensor. Development of such nanoporous structure enables the miniaturization of high-performance electrochemical biosensors for point-of-care diagnostics or environmental ﬁeld testing. & 2012 Elsevier B.V. All rights reserved. 1. Introduction Lab-on-a-chip technology has attracted researchers from dif- ferent disciplines to explore this technology for a wide range of applications (Chin et al., 2012; Ducre ´e et al., 2007; Huang et al., 2012; Madou et al., 2006; Whitesides, 2006). Enzymatic electro- chemistry is one of the most applied detection techniques used in lab-on-a-chip devices to miniaturize chemical and biological analysis processes due to its low cost, high sensitivity, moderate power requirements, and prominent compatibility with micro- fabrication technologies (Mir et al., 2009; Vandaveer et al., 2004; Wang et al., 2001). Although the miniaturization will not alter the mechanism of biochemical reactions on electrodes, it will change ﬂuid mechanics, molecular diffusion and surface to volume ratio in micro scale channels and will require the modiﬁcation of bulk electrochemical biosensors. All types of enzymatic electrochemical biosensors, electrodes (working electrode, reference electrode and counter electrode) perform as transducers, which convert information of a speciﬁed amount of biological analyte into an electrical signal. The bior- ecognition phenomenon and the redox reaction occur on the surface of the working electrode where the catalyst layer, the enzyme layer and the semi-permeable layer are superimposed. Therefore, the working electrodes employed in microﬂuidic chips deserve considerable attention. To ﬁt the electrode in the detec- tion chamber and to minimize the disturbance to ﬂuid movement, the thickness of the embedded planar electrode must be reduced to nanoscale and the surface area must be as small as possible. However, the miniaturization results in several implications require further improvement. For example, the miniaturization impairs the signal-to-noise ratio and the nanoscale thickness of the metal layer limits the choice of surface functionalization for subsequent coating. Because of the increasing demand for apply- ing electrochemical sensing technique on Lab-on-a-chip devices, the electrochemical sensing surface is expected to be thinner and possess a higher signal-to-noise ratio. Efforts have been made to enhance the sensing sensitivity through artiﬁcially enlarging the surface area of working electrodes. Recently, porous materials prepared by sol–gel methods have been developed to immobilize enzymes (Carturan et al., 1998; Liu et al., 2000). Nanostructured materials have also attracted much attention due to their inherently large surface area. For example, carbon nanotubes, which were randomly or uniformly immobilized with enzymes, were used in the conﬁguration of biosensors (Wang, 2005). Metal nanoparticles and nanowires were also prevalent selections for substrate materials (Hrapovic et al., 2004). Contents lists available at SciVerse ScienceDirect journal homepage: www.elsevier.com/locate/bios Biosensors and Bioelectronics 0956-5663/$ - see front matter & 2012 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.bios.2012.10.003 n Corresponding author. E-mail address: jyang@eng.uwo.ca (J. Yang). Biosensors and Bioelectronics 42 (2013) 5–11