Development of an impedimetric biosensor for lung cancer detection.

The promise of biosensors offering attractive features has kept the field active and growing. The aim is often to develop a device which is sensitive, specific, rapid, portable, cheap, and with the ability to capture an analyte in different matrices without cross-reactivity. The challenges increase when the aim is simultaneous multi-analyte detection on a single platform, without the need for complex procedures and expensive instruments. Such a system is invaluable in many clinical settings, where disease diagnosis and progression are multifactorial. Cancer is a good example of complex diagnosis requirements. This project is aimed at the development of biosensing platform to overcome the aforementioned problems and allow direct, simple analysis with the minimum of sample pre-treatment. The early detection of lung cancer has been chosen as there is no commercial biosensor available for detection of this disease and for lung cancer diagnosis multi-analyte recognition is necessary. This project presents the development of impedimetric and magnetic sensing platform for early detection of lung cancer via detecting the neuron-specific enolase (NSE) and carcinoembryonic antigen (CEA) which are known as potential lung cancer biomarkers. The sensing platform developed here comprises of magnetic manipulation, screen printed electrode (SPE), and magnetic nanobeads. The magnetic nanobeads (MBs) were functionalised with antibodies to fish the analyte from the sample, and to move them over the sensing area. Moreover, magnetic nanobeads were used to increase the chance of antigen-antibody complex formation. After cleaning the surface of electrodes with 50 mM KOH in 25% H₂O₂ (for 10 minutes), immunosensors were developed by immobilising the antibodies on the gold working electrode of SPEs through formation of self-assembled monolayer (SAM layer) due to its simplicity, stability, well-organised structure and low background noise. The optimised NSE immunosensor with 10 µg/ml of 10-7937 antibody was successfully tested to measure various concentrations of NSE protein (0 – 100 ng/ml) in both PBS buffer and 100 % serum using functionalised MBs with 2.4 mg/ml of 10-7938 antibody. The optimised sensor achieved detection limit of 0.18 ng/ml (R²= 0.9848) in buffer and 0.52 ng/ml (R² = 0.9977) in 100 % serum with via EIS with the use of 10 mM potassium ferri/ferrocyanide as a redox probe. The impedimetric CEA immunosensor was developed and optimised by use of 20 µg/ml of 12-140-01 antibody and was used to measure analyte with functionalised MBs with 2.4 mg/ml of 12-140-10 antibody. The CEA immunosensor was also able to measure various CEA concentrations (0 – 100 ng/ml) in both PBS buffer and 100 % serum in presence of 10 mM potassium ferri/ferrocyanide. The sensor achieved low limit of detection as 0.26 ng/ml (R² = 0.9924) and 0.76 ng/ml (R²= 0.9839) for CEA detection in buffer and 100 % serum, respectively. In conclusion, both immunosensors developed here, using EIS and the magnetic sensing platform, were capable of detecting their corresponding biomarkers in serum in relatively short time (40 minutes) and in the appropriate concentration range, as serum concentrations higher than 12.5 ng/ml and 7 ng/ml for NSE and CEA respectively can indicate presence of cancer. To the best of our knowledge, no one has reported a use of magnetic platform as the one developed in this thesis.