Label-Free Protein Detection on Graphene Oxide Surface


Kazuaki Furukawa and Yuko Ueno*
Materials Science Laboratory, *NTT Microsystem Integration Laboratories

   Graphene oxide (GO) is an analogue of an oxidized form of graphene, which contains a number of C-O bonds generated by the oxidization of C=C double bonds. GO holds an atomically thin sheet-like structure like graphene, but at the same time it shows many different features from graphene. For instance GO is an insulator, water-dispersive material and efficient fluorescence quencher. We designed and built the label-free protein recognition system on the GO surface using these unique properties [1].
   We used GO fixed on a solid surface. The GO surface was modified using pyrene linker to sp2 domains left in GO and the thrombin aptamer with a probe dye (FAM) bonded at the other terminus (Fig. 1, top). Here the aptamer is a single strand DNA that forms a complex with a specific target molecule. In this study, we used the aptamer for thrombin, an important protein for blood clotting, to demonstrate the idea.
   The recognition mechanism is schematically shown in Fig. 1. Fluorescence from the dye is quenched by the GO at the initial stage when the modified molecules adsorbed on the GO surface with the strong interaction between single strand DNA and GO. The fluorescence recovers when the aptamer recognizes and forms a complex with thrombin (Fig. 1, bottom). This is because the dye is separated from the GO surface by the recognition. The validity of the system was confirmed using a single piece of GO. Fluorescence microscope observations showed that the fluorescence recovers only on the GO surface after the thrombin addition (Fig. 2, top). The thickness of GO was increased about 2.9 nm upon the thrombin recognition, which was traced by atomic force microscope observations for the same GO piece (Fig. 2, bottom). The experiments demonstrated the successful label-free detection of thrombin. Our system is advantageous for the operation in microfluidics because the GO piece is firmly fixed on the solid surface. With an increase variety of detectable proteins by choosing different aptamers, it provides a common platform of GO aptasensors.

[1] K. Furukawa et al., J. Mater. Chem. B 1 (2013) 1119.

Fig. 1. Schematic drawing of aptamer-modified graphene oxide surface and its molecular detection mechanism.
Fig. 2. Fluorescence microscope (top) and atomic force microscope (bottom) images of a GO piece before and after thrombin addition.