3-Dimensional Graphene based Electrochemical Biosensor for Extracellular Vesicle Detection

Over expression of Extracellular vesicles is a universal effect of the presence of a carcinoma in the human body. Biomarkers such as Tspan-28 provide a clear target, on the surface of EVs, to understand the stage and progression of cancer in a patient. Measuring EV levels could indicate the presence and stage of a cancer. With minute deviations in concentrations of such a biological marker, EVs can be used as a target for tracking and diagnosing cancer at an early stage. Detecting cancer at an early stage is an extremely important task, as early detection can lead to a more favourable prognosis compared to later diagnosis. To address this task, we used a commercially bought 3D porous graphene foam (3D-PGF) platform with exceptional electrical conductivity and structural qualities. Utilizing the malleable surface for modification, we were able to produce a modified 3D-PGF structure of 1-pyrenebutyric acid N[1]Hydroxysuccinimide ester (PYB-NHS) upon which specific antibodies to Tspan-28 were immobilized. This provided a structurally and electrochemically stable platform for the detection of the Tspan-28 protein expressing EVs and pure Tspan-28 in human serum. The capture of EVs to the modified electrode platform was analysed with electrochemical impedance spectroscopy (EIS) and using a label-free ferri/ferro redox reaction method to analyse the surface change after protein detection. The 3D-PGF platform provides an ultrasensitive detection range from 0.1 fg/ml to 1000 fg/ml with pure Tspan-28 protein and a linear range of 10^2 -10^5 EVs /mL with all samples in a 10:1 PBS human serum
mixture for realistic real-world results. The LOD for EV detection was calculated at ∼24 EVs /mL (n = 3), which is an unprecedented level of detection for label-free electrochemical sensors and can provide the ultrasensitive levels needed for early cancer detection.