Abstract: Understanding molecular mechanisms of diseases and manipulating them to achieve cure constitutes one of the "holy Grail" of biomedical science. Infectious (and parasitic) diseases are responsible for ~30% of premature deaths, especially in low- and middle-income countries. Studies of the pathogenic agents require high-resolution and high-sensitivity microscopy and spectroscopy equipment, ideally capable of detecting single molecules of interest (i.e., DNA bases, proteins on viral capsids, etc.). Scanning near-field optical microscopy (SNOM) combined with near-field nano-spectroscopy- Tip-Enhanced Raman spectroscopy (TERS) is the best technology for visualization of such nanoscale details. However, the spatial resolution and chemical sensitivity levels are currently insufficient to detect and identify single molecules in an ambient environment (air or liquid). Current methods for the identification of infectious agents rely on traditional methods that include growing of bacteria and fungi, isolation of viruses in cell culture, and then identification of the agent biochemically, antigenically, or genetically (which require the availability of specific antibodies for ELISA tests and specific DNA primers for PCR). These methods make the identification of newly emerging pathogens particularly difficult. New approaches to diagnostics are overdue. Actoprobe LLC proposes such an approach: to develop, validate, and commercialize a novel class of probes for Scanning Probe Microscopy (SPM) based on silicon nitride photonics with a diode laser, a pulse compressor, and a detector, all inside the SPM probe. The ultimate goal is to dramatically (>10X) increase SNOM's sensitivity and spatial resolution for rapid identification of infectious agents. With its strong research experience and expertise in TERS and SNOM, Actoprobe is in an ideal position to address this technical challenge. This new TERS tool's capabilities will allow studying the biochemical components, morphology, and nanomechanical structures of viruses, bacteria, and fungi at the single-molecule level, substantially improving early infectious disease detection and identification. We expect the proposed technology will enable substantially improved early pathogenic disease agent detection and lead to a marketable, cost-effective diagnostic/identification tool. Our primary customers will be medical laboratories and clinical research institutions. Phase I will prove the technical feasibility of developing the ultrasensitive optical SPM probes and testing these probes on several non-pathogenic viruses.

Narrative New infectious disease diagnostic approaches are required to improve diagnostic speed, reliability, and detection sensitivity, ideally for label-free disease-causing agent detection directly from patient samples. Characterization of single DNA and RNA at the single-molecule level, however, remains unattainable due to the lack of applicable technologies that can reach single-molecule resolution and chemical sensitivity. This project will develop a novel class of scanning probe microscopy near-field optical probes that will provide new and enhanced research capabilities for studying biochemical components of viruses, bacteria, fungi, and other microorganisms and their morphology, ultrastructure, and nanomechanical properties, with the ultimate goal to enable very long fragments of DNA /RNA sequencing with no previous information about the organisms to do early infection detection and identification.