This proposal targets the problems of the Debye length in using FET for detection of biomolecules. Since the distance of detection is limited to 10 nm or less, antibodies have not been used to target biomolecules using FET sensors since antibody length is beyond the Debye length. The investigators have taken the approach of using just the targeting antibody fragment for HEMT. To achieve this goal, the approaches taken include: (1) to identify antibody fragments (scFvs and VHH) that recognize MIG (analyte receptors) and engineer selected scFv and VHH thereof for site specific conjugation to HEMT sensing surfaces; (2) to characterize specificity and sensitivity of MIG-detecting HEMTs decorated with scFvs and VHHs and the chemical and electronic properties of sensing interfaces; (3) to use a control gate or an optimized structure on scaled nano-device for sensitivity enhancement and (4) to establish theoretical models and perform simulations, coupled with experimental results, for fundamental understanding.
NSF 0756594 aided the realization of an immunologically-modified field effect transistor (immunoFET) sensor consisting of semiconductor transistor (an AlGaN/GaN heterojunction field effect transistor) modified to detect specific proteins in physiologic buffers. The scientific impact of the realization is potentially large: immunoHFET assays are a potential low labor, real-time replacement for ELISA. ImmunoHFET assay directly measures analyte properties (charge, field strength) and do not require secondary reagents, unlike ELISA, Western analysis or most immunoassays. ImmunoFET sensors have protein analyte-specific antibodies deployed on their sensing surfaces. Since proteins possess an inherent charge/electrical field, protein binding to the antibodies on the sensing channel brings analyte charge/electrical field proximal to the sensor surface. This, in turn, induces detectable changes in transistor electrical properties. We monitor change in transistor current and other electrical properties, with analyte binding, and current parameters can be related to sample protein concentration by a standard curve for individual analytes. Knowledge of specific protein concentrations (in this case, concentrations of immunomodulatory proteins) is of critical importance in both research and clinical settings. The development of an immunoFET exploits not only the robust capabilities of the semiconductor industry, but also the rapid and sensitive responses of these electrical devices. The result is accurate, reliable protein sensing in real-time. It should be noted that the utility of immunoHFETs is not limited to immunomodulatory cytokines/chemokines. We have successfully detected multiple unrelated proteins, including antibodies, streptavidin, avidin and albumin as well. ImmunoHFETs thus represent a technology for protein detection and quantification applicable to many, if not all, classes of polypeptides. Since our first successful detection of protein in a physiologic salt environment (Gupta et al., 2008), the focus of the project has been on three interrelated areas. The first was to obtain a more thorough understanding of the sensor interface where protein binding occurs in both immunoHFETs and bioHFETs (Bhushan et al., 2009, Gupta et al., 2011); the second was to improve the sensitivity of the device to allow for the detection of proteins across all clinically reported levels for our proteins of interest (Nicholson et al., 2011); and third was the detection of native, unlabeled proteins by genuine immunoFETs (Casal et al., 2012). More recently, our project’s focus has moved towards a partnership with the Ohio State University’s Division of Transplantation surgery. This collaboration aims to use the protein detection capabilities of our immunoHFET sensor to detect the levels of specific proteins linked to transplant rejection. The goal is to provide real-time, point-of-care devices for determining the rejection status of patients’ grafts and guiding their care appropriately (through adjustment of immunosuppression regimens). ImmunoHFET sensors offer the opportunity to detect rejection early, allowing time for clinical intervention prior to irreversible damage to the graft. Given the ability of immunoHFET sensors to detect a broad range of unrelated proteins, with appropriately configured immunoHFETs, the technology offers the opportunity to detect protein analytes in multiple environments. The technology should be applicable to any disease state wherein protein concentration is an issue of importance, including multiple metabolic disease and infectious disease. Reflecting the commercial/clinical potential of immunoHFETs operated under physiologic conditions, NSF 00756594 has laid the groundwork to found an immunoHFET company (Sensetronics) and for submission of an application for a Small Business Innovation Research (SBIR) grant. SBIR funding will be used to develop Sensetronics commercial products from the research conducted under NSF 0756594 for clinical use.
