Electrochemical endotoxin sensors based on TLR4/MD-2 complexes immobilized on gold electrodes

Abstract

Even low concentrations of endotoxins can be life-threatening. As such, continuous effort has been directed toward the development of sensitive and specific endotoxin detection systems. In this paper, we report the design and fabrication of a new electrochemical endotoxin sensor based on a human recombinant toll-like receptor 4 (rhTLR4) and myeloid differentiation-2 (MD-2) complex. The rhTLR4/MD-2 complex, which specifically binds to endotoxin, was immobilized on gold electrodes through a self-assembled monolayer (SAM) technique involving the use of dithiobis(succinimidyl undecanoate) (DSU). The surface topography of the electrodes at each fabrication stage was characterized with a nanosurface profiler and atomic force microscope (AFM). The electrochemical signals generated from interactions between the rhTLR4/MD-2 complex and the endotoxin were characterized by cyclic voltammetry (CV) and differential pulse voltammetry (DPV). A linear relationship between the peak current and endotoxin concentration was obtained in the range of 0.0005 to 5 EU/mL with a correlation coefficient (R(2)) of 0.978. The estimated limit of detection (LOD) was fairly low, 0.0002 EU/mL The rhTLR4/MD-2 based sensors exhibited no current responses to dipalmitoylphosphatidylcholine (DPPC) bearing two lipid chains, which is structurally similar to endotoxin, indicating the high specificity of the sensors to endotoxin. (C) 2011 Elsevier B.V. All rights reserved.