Solid-state nanopores have been widely employed in different sensing applications from Coulter counter to DNA sequencing devices. These versatile sensors can detect analytes of different size ranging from single molecules to bacteria. However, most reported applications of nanopores are qualitative, and their physical size and geometry are often not suitable for in situ measurements in microenvironments. This proposal is aimed at developing a new biosensor platform for the detection of antibodies and protein biomarkers based on the use of nanometer-sized pipettes. Like a nanopore, a pipette can sense the analyte that enters its aperture and partially blocks the ion current flowing through it. However, a nanopipette offers several important advantages including the ease of fabrication, small physical size, and needle-like geometry that makes it suitable as a probe for scanning electrochemical microscopy (SECM). The translocation of metal nanoparticles will be used as a model system to establish the principles of resistive-pulse sensing with nanopipettes. By thoroughly characterizing the pipette size and geometry, we will demonstrate the quantitative relationship between the frequency of the recorded single particle events and the diffusion flux of particles to the orifice. This will enable quantitative determination of analyte species in solution. Following proof-of-concept experiments with bare metal particles, we will work on the detection and quantitation of antibodies (e.g., antipeanut antibody) attached to nanoparticles and cancer biomarkers (e.g., prostate specific antigen, PSA). Overall, the proposed work will result in the development of a new biosensing platform for quantitative analysis at the level of single biomolecules or single nanoparticles. This potentially transformative concept can cause the paradigm shift in bioelectroanalytical chemistry. Broader Impacts The proposed approach to biosensing can enable accurate diagnostics of allergies, cancer and other diseases based on the detection of low concentrations of antibodies and protein biomarkers. It can find other applications in biomedical testing including DNA sequencing. The requested support is mostly for graduate and postdoctoral stipends. The graduate students and postdoctoral fellows involved in this project will get multidisciplinary research experience in sensors, electrochemistry, bioanalytical chemistry and nanoscience. The results of this research will be broadly disseminated through publications and professional presentations. The requested funds will enable us to contribute to ongoing CUNY efforts to recruit underrepresented minority STEM students and participate in its diversity initiatives. This project will provide research opportunities to undergraduates and high school students.
This project resulted in the development of nanometer-sized pipettes as resistive-pulse sensors. Pulled quartz pipettes were used for label-free detection of peptide-modified particles and antibodies attached to them, as well as a cancer biomarker (prostate specific antigen, PSA). The possibility of probing mixtures of different analytes was also demonstrated. We improved the procedures for characterizing nanopipette geometry and theoretical treatment for the interpretation of the resistive-pulse data and compared the obtained particle size distributions to the results produced by other techniques. The experimental protocol was developed for fabricating open carbon pipettes with different size and geometry by coating the inside of pulled quartz capillaries with a nanometer-thick carbon film. The extent of ion current rectification in a carbon pipette depends on the potential applied to the carbon layer, which determines the surface charge at the pipette wall. Thus, such a pipette can work as a tunable resistive-pulse sensor whose properties can be adjusted to detect a specific analyte without chemical modification of the inner pipette wall. The very small attainable diameter of a carbon pipette tip (<20 nm) makes it potentially useful for local electrochemical and resistive-pulse measurements. This project provided opportunities for multidisciplinary research to several graduate students. They were trained in electrochemistry, bioanalytical chemistry and nanoscience. These students gave oral and poster presentations at national and international meetings and participated in preparation of research papers. They also visited the laboratories of our collaborators and performed joint experiments with their groups. One student obtained her PhD degree and started postdoctoral studies. A visiting graduate student from China has spent two years in the PI's lab, thus establishing a new international collaboration.