Graphene field-effect transistor biosensor for detection of biotin with ultrahigh sensitivity and specificity

Abstract

Because avidin and biotin molecules exhibit the most specific and strongest non-covalent interaction, avidin-biotin technology is widely used in ELISA (enzyme-linked immunosorbent assay) kits for the detection of different bio-macromolecules linked to different diseases including cancer and influenza. Combining the outstanding electrical conductivity (200,000 cm²V^-1s^-1) of graphene with the unique avidin and biotin interaction, we demonstrate a novel graphene field-effect transistor (GFET) biosensor for the quantitative detection of bio-macromolecules. The GFET consists of six pairs of interdigital Cr/Au electrodes supported on Si/SiO₂ substrate with an avidin immobilized single layer graphene channel as the sensing platform. By monitoring the real time current change upon the addition of biotin solution in bovine serum albumin (BSA) in the silicone pool preformed onto the GFET, the lowest detectable biotin concentration is estimated to be 90 fg/ml (0.37 pM). The specificity of the GFET is confirmed both by controlled and real sample measurements. From the magnitude of current change upon the addition of different concentrations of biotin solutions, the dissociation constant K_d is estimated to be 1.6 × 10^-11 M. Since biotin is capable of conjugating with proteins, nucleotides and other bio-macromolecules without altering their properties, the present GFET sensor with its ultra-high sensitivity (0.37 pM) and specificity can be tailored to the rapid point-of-care detection of different types of desired biomolecules at very low concentration level through biotinylation as well as the exogenous biotin in blood serum.

Keywords: Avidin; Biosensor; Biotin; Clinical diagnosis; Field-effect transistor; Graphene.

Fig. 1

(a) The full-scale optical image of the interdigital electrodes fabricated on a SiO₂/Si substrate. (b) The 3D image showing the height of the interdigital electrode and substrate.

Fig. 2

(a) Schematic illustration of the graphene biosensor fabricated on SiO₂/Si substrate. Graphene was carefully transferred to cover all six pairs of interdigital electrodes. (b) The typical Raman spectrum of the graphene transferred onto the interdigital electrodes.

Fig. 3

The schematic illustration of the stepwise modification of graphene with avidin molecules followed by the binding of biotin molecules.

Fig. 4

(a) Raman spectra of the transferred graphene after modification with PBASE for the different times indicated (1 h, 2 h, 4 h, 6 h, 8 h). (b) The change of 2D/G ratio of the Raman spectra shown in (a). (c) The I–V curves of the bare graphene and after 4 h modification with PBASE. (d) The high-resolution N 1s XPS spectra of the clean, PBASE modified and avidin modified graphene.

Fig. 5

(a) The 2-terminal real-time test system. A silicone pool is constructed onto the graphene transferred on interdigital electrodes fabricated on SiO₂/Si substrate. (b) The real time current (Ids) upon addition of different concentrations of biotin molecules. (c) Langmuir fitting to the changes of Ids at different concentration of biotin. (d) The real-time current (Ids) upon the addition of PBS, BSA, biotin, and BSA at different times.