High-Density Immobilization of Tobacco Mosaic Virus Nanotubes As Enzyme Nanocarriers Onto Field-Effect Biosensor Structures

Abstract

Introduction Viruses are not only infectious agents, they are also known as promising functional building blocks for application in nano- and biotechnologies. The tobacco mosaic virus (TMV) was the first studied plant virus and is widely distributed; it infects vegetables, like tomato, bell pepper, beans and other members of the family Solanaceae, while it is totally harmless for mammals [1]. It is one of the most studied plant viruses and its genome is completely sequenced, whereby genetical and chemical modification is easy. TMV has a nanotube-like shape with a length of 300 nm, an outer diameter of 18 nm and an inner diameter of 4 nm. Since it possesses a high chemical and physical robustness, it can be integrated with different electronic transducers for bio- and chemical sensing applications [2].Recently, we presented a TMV-based amperometric glucose biosensor [3] and a potentiometric penicillin biosensor [4]. TMV was used as enzyme nanocarrier for the enzyme glucose oxidase and penicillinase, respectively [3,4]. The sensitivity and detection limit of these biosensors, among others, depend on the density of TMVs on the sensor surface. The surface density of the immobilized TMVs is strongly influenced by the electrostatic interactions between the charged TMVs and sensor surface as well as by the inter-TMV-nanotubes repulsion, which could be changed by varying the pH value and the ionic strength of the TMV solution. In this study, we investigated an impact of the pH value and ionic strength of the TMV solution on the surface density of TMV nanotubes immobilized onto Ta2O5-gate capacitive field-effect electrolyte-insulator-semiconductor (EIS) sensors. Materials and Methods TMV particles modified with biotin-linker molecules (TMVBio), which serve as binding sites for streptavidin-conjugated enzymes, have been immobilized onto capacitive field-effect EIS sensors with Ta2O5 as transducer layer, as shown in Fig. 1. Immobilization was performed from TMVBiosolutions with different values of pH between pH 3.0 and pH 9.5 and ionic strength between 0.1 mM and 750 mM. The sensors have been electrochemically characterized before and after TMVBio immobilization by capacitance-voltage- and constant-capacitance methods, respectively. In addition, the density of the immobilized TMVs and morphology of the sensor surface has been investigated by means of scanning electron microscopy (SEM). Results and Conclusions The TMVBio density on the Ta2O5 sensor surface was influenced by varying the pH value (as exemplarily shown in Fig. 1) and ionic strength (not shown) of the TMVBio solution. The amplitude of the field-effect sensor signal correlates well with the density of the immobilized TMVBionanotubes. Thus, optimized conditions for the high-density immobilization of TMVBio nanotubes onto the Ta2O5 surface and thereby enhanced biosensing have been found. Details of the experiments and the obtained results will be presented and discussed.Figure 1: Measurement set-up with schematic layer structure of the capacitive EIS sensor modified with negatively charged TMVBio particles (a). SEM images of the Ta2O5-sensor surface modified with TMVBio nanotubes at pH 4.5 (b) and pH 7.0 (c), respectively. Acknowledgements The authors like to thank Dr. Claudia Koch and Rebecca Hummel, Stuttgart, for scientific and technical support.