Biosensors for detection of colorectal cancer

Morbidity and mortality rates of colorectal cancer (CRC) remain among the highest in cancer cases due to late detection. Conventional diagnostics are complicated, often invasive and time-consuming with variable sensitivity and specificity. Carcinoembryonic antigen (CEA) is the only validated blood biomarker routinely used for prognostic screening of advanced CRC. Therefore, development of a highly sensitive, specific and rapid diagnosis device, such as a biosensors is needed. Most biosensors developed currently are antibody-based which restricts their commercialisation due to stability, cost and variability batch-to-batch. The main aim of this study was to develop impedimetric biosensors that are inexpensive and with simple fabrication using anti-CEA Affimers as novel bioreceptors. Initially, the anti-CEA Affimers were selected from a phage display library and characterized for specificity binding against CEA. Ultimately, anti-CEA Affimer-based biosensors were fabricated on DropSens screen printed gold electrodes coated with a novel non-conducting polymer layer, polyoctopamine (POct), for CEA detection. Electrochemical characterisation of developed biosensors was performed via cyclic voltammetry, electrochemical impedance spectroscopy and protein blotting. Newly constructed CEA immunosensors with anti-CEA IgG covalently immobilised onto the POct coated electrode surface were developed in parallel as a comparison. A highly sensitive and specific anti-CEA Affimer-based biosensor was successfully developed that is superior to the monoclonal based CEA immunosensor when both were interrogated in buffer or in diluted human serum. A polyclonal based CEA immunosensor showed equivalent sensitivity to the Affimer-based biosensors with limit of detection (LOD) at 1 fM and wide dynamic range (1 fM – 100 nM). This LOD was significantly lower than the basal clinical levels of 25 pM (i.e.  5 ng/ml). Overall, this label-free Affimer-based biosensor has strong potential to be developed as point-of-care device for CEA detection as it required small sample volumes (10 μl) and had rapid response time (5 min). Additionally, the newly discovered polyoctopamine polymer base layer has proved its generic application in immobilising different types of bioreceptors.