2025 Theses Doctoral

Affinity Nanobiosensors toward Clinical Monitoring of Biomarkers

Dai, Wenting

Affinity biosensors are crucial in clinical settings for their ability to provide rapid, accurate and sensitive detection of analytes, enabling early diagnosis and monitoring of diseases. These biosensors operate on the principle of high-affinity binding between recognition receptors and the target analyte to cause a physicochemical change which is converted by a transducer into a measured signals to obtain the analyte concentration. Current commercial affinity biosensors, utilizing optical, electrochemical and mass-based transduction methods, offer high sensitivity and specificity of analyte measurements for clinical applications, but face limitations such as high costs and bulkiness for optical biosensors, difficulty in detecting small molecules for mass-based biosensors, and complexity and label degradation issues for electrochemical biosensors.

2D nanomaterial-based aptameric nanosensors, which utilize 2D nanomaterials as transduction materials and aptamers as receptors to specifically recognize target analytes, hold the potential to address these limitations of traditional commercial biosensors. However, the nanosenors still face challenges for practical applications in clinical settings, such as nonspecific binding in physiological media, the necessity for individual sensor calibration, and limitations in current 2D nanomaterial properties and performance degradation over time. This thesis focuses on developing 2D-nanomaterial based aptameric FET nanosensors to enhance their adaptability toward practical clinical applications.

We first present the optimization of surface modification of aptameric graphene nanosensors for measurement of biomarkers in undiluted physiological media. In these sensors, graphene surfaces are coated with a polyethylene glycol (PEG) nanolayer to minimize nonspecific adsorption of matrix molecules. We perform a systematic study of the aptamer and PEG attachment schemes and parameters, including the impact of the serial versus parallel PEG and aptamer attachment scheme, PEG molecular weight and surface density, and aptamer surface density on the sensor behavior, and then use the understanding from this parametric study to identify optimal surface modification of the nanosensor to enable sensitive and specific biomarker measurements in undiluted physiological media.

We then present a calibration-free method that exploits kinetic measurements to enable the calibration-free operation of aptameric graphene nanosensors. In our kinetically based calibration-free method, a time constant estimated based on the aptamer-analyte kinetic binding process is used to determine the analyte concentration. The time constant depends on the to-be-measured analyte concentration and reaction rate constants 𝑘_on and 𝑘_off, respectively, which is unaffected by variations in measurement conditions and sensor properties, thereby allowing devices functionalized with the same aptamer to be used without individual calibration once one particular device is calibrated, and enabling accurate analyte concentration determination independent of variations in parameters.

Next, we present an affinity nanosensor that uses Ni₃(HITP)₂ metal-organic framework (MOF) as the conducting channel which is functionalized with an aptamer for specific biomarker recognition. Binding between the aptamer and the target biomarker induces a change in the carrier density in the MOF and resulting in measurable changes in FET characteristics for determination of the biomarker concentration. 𝘐𝘯-𝘴𝘪𝘵𝘶 synthesis of the MOF enhances the adhesion between the MOF film and electrodes to reduce noise during sensor operations, while also allowing devices to be potentially mass-produceable at reduced cost. Thanks to the porous structure of the MOF, the aptamer surface density on the MOF is optimized to enhance sensitivity for biomarker measurements.

We finally present the preliminary development of aptameric graphene nanosensors for accurate and stable extended-time measurement of analyte. We perform a systematic study of the stability of the nanosensor including the graphene, aptamers and aptamer attachment, to identify the cause of performance degradation, and use the understanding to develop a differential nanosensor functionalized the aptamer through covalent reaction and implement graphene compensation method to achieve the accurate and stable extended-time analyte measurement.