[Frontiers in Bioscience 10, 2518-2526, September 1, 2005] 2518 IMPEDANCE LABELLESS DETECTION-BASED POLYPYRROLE PROTEIN BIOSENSOR C. M. Li 1 , Wei Chen 1 , Xu Yang 1,2 , C. Q. Sun 3 , Crystal Gao 4 , Z. X. Zheng 4 and Jaymie Sawyer 4 1 School of Chemical & Biomedical Engineering, Nanyang Technological University, Nanyang Avenue, Singapore 639798. 2 College of Life Science, Central China Normal University, Wuhan 430079, China. 3 School of Electrical and Electronic Engineering, Nanyang Technological University, Nanyang Avenue, Singapore 639798. 4 Motorola Life Science, Tempe, Arizona 85287, USA. TABLE OF CONTENTS 1. Abstract 2. Introduction 3. Materials and Methods 3.1. Materials and solutions 3.2. Apparatus 3.3. Experimental procedure 4. Result and discussion 4.1. Impedance detection of Ab-Ag (antibody-antigen) interactions 4.2. Data processing for S/N (signal to noise) ratio improvement 4.3. Specificity of impedance labelless detection of proteins 4.4. Nonspecific binding effect in serum sample 5. Conclusions 6. Acknowledgment 7. References 1. ABSTRACT A simple and sensitive electrochemical immunosensor with impedance labelless detection and novel data processing method was investigated. One-step polymerization was used to electrochemically deposit an antibody impregnated polypyrrole film on a glassy carbon electrode surface for the immunosensor. Impedance measurements provided a labelless or reporterless method to detect antibody (Ab)-antigen (Ag) interactions. Dimensionless analysis was employed to successfully process the measured impedance data. Since the method derived unit impedance change to eliminate or reduce the variation of the bulk electronic properties of Ab/polypyrrole films, the signal to noise ratio (S/N) was significantly improved for high sensitivity and specificity. Nonspecific binding effect was studied by array electrode chips and was found out that the polypyrrole electrode without antibody attachment had much stronger nonspecific binding effect than the Ab/polypyrrole electrode; incubation followed by thoroughly washing significantly reduced the nonspecific interference. 10 pg/ml detection limit and superior specificity were achieved by the method, demonstrating a highly sensitive labelless immunosensor in comparison with the detection limit of ng -µg/ml for the reported polypyrrole based immunosensors. The electrochemical immunosensors presented in this paper, due to its simplicity, low cost, high sensitivity and superior specificity, could be an invaluable tool for clinical diagnostics and could have potential applications in drug discovery, environmental and food analysis.