SBIR-STTR Award

This Phase I proposal aims to develop and demonstrate the feasibility of Fluorescence Enhanced Photothermal Infrared (FE-PTIR) imaging and spectroscopy. The proposed FE-PTIR will use fluorescence microscopy to map the distribution of fluorescently labeled regions of cells and tissue and then provide chemical structural analysis of the labeled regions using photothermal infrared spectroscopy. Fluorescence microscopy is a cornerstone technique in biological research, allowing sensitive and highly specific mapping of target biomolecules within cells and tissue, but it does not provide information about the chemical structure of those molecules. Infrared spectroscopy can provide rich analysis of chemical structure and has been used in life sciences research to study tissue classification, drug/tissue interaction, neurodegenerative diseases, cancer research and other areas. Conventional infrared spectroscopy, however, has a fundamental limit on its spatial resolution (i.e. roughly how small an object it can analyze) of around 10 micrometers, similar to the size of an average biological cell. Thus conventional infrared spectroscopy has been extremely limited for many biomedical applications where the structures of interest are smaller than the size of a cell. The proposed FE-PTIR technique will overcome the limitation of both fluorescence microscopy and infrared spectroscopy to provide highly specific mapping of target biomolecules along with chemical structural analysis of those molecules, both with the same spatial resolution as fluorescence microscopy. This project will achieve this breakthrough by using a novel form of optical photothermal infrared spectroscopy to measure infrared spectra of fluorescently labeled regions of a sample. Specifically, the FE-PTIR technique will illuminate a sample with an infrared laser source that can be tuned to excite molecular vibrations a sample of interest. A separate ultraviolet/visible light source will be used for two jobs: (1) to excite fluorescent emission in fluorescently labeled regions of the sample; and (2) measure a localized heating resulting from absorption of infrared radiation. By measuring the intensity of fluorescent light emitted from different regions of the sample, it is possible to map the distribution of fluorescently labeled biomolecules. Then by measuring subtle changes in the amount of UV/visible light collected from the sample resulting from the local IR-induced heating, it is possible to generate infrared absorption spectra of the same locations and with the same spatial resolution. The infrared absorption spectrum can then be used to analyze the chemical structure of the fluorescently labeled regions of the sample. This project is well aligned with NIH goals as it incorporates several key thrusts of the National Institute of Biomedical Imaging and Bioengineering, including optical imaging and spectroscopy, IR imaging, confocal microscopy, and multimodal imaging. FE-PTIR will be extremely useful for example in localizing specific proteins with fluorescence microscopy and then analyzing using photothermal IR spectroscopy to analyze their structure, for example how the protein is folded. Protein misfolding is a root cause of many neurodegenerative diseases (e.g. Alzheimer's) and FE-PTIR will offer new insights. Demonstrating the FE-PTIR technology will enable a new multimodal microscope with sub-cellular resolution that will offer profound benefits for biomedical research including neurodegenerative diseases and antimicrobial resistance research.

Public Health Relevance Statement:
Project narrative Fluorescence microscopy is a cornerstone technique in biological research that maps the distribution of fluorescently labeled biomolecules but does not provide information on their chemical structure. Infrared spectroscopy provides detailed chemical structural analysis on biological materials, but fundamental resolution limits have constrained its application in biology. This SBIR project will overcome these limits to enable both mapping and chemical analysis of target biomolecules with sub-micron spatial resolution, thus enabling research to provide fundamental insights into neurodegenerative diseases and antimicrobial resistance.

Project Terms:
absorption; Alzheimer's Disease; AD dementia; Alzheimer; Alzheimer Type Dementia; Alzheimer disease; Alzheimer sclerosis; Alzheimer syndrome; Alzheimer's; Alzheimer's disease dementia; Alzheimers Dementia; Alzheimers disease; Primary Senile Degenerative Dementia; dementia of the Alzheimer type; primary degenerative dementia; senile dementia of the Alzheimer type; Biocompatible Materials; Biomaterials; biological material; Biological Sciences; Biologic Sciences; Bioscience; Life Sciences; Biology; Biomedical Research; Boston; Cells; Cell Body; Classification; Systematics; Pharmaceutical Preparations; Drugs; Medication; Pharmaceutic Preparations; drug/agent; Fluorescence; Goals; Heating; Infrared Rays; infrared radiation; Lasers; Laser Electromagnetic; Laser Radiation; Light; Photoradiation; Maps; Microscopy; Fluorescence Microscopy; Fluorescence Light Microscopy; United States National Institutes of Health; NIH; National Institutes of Health; Occupations; Jobs; Professional Positions; Optics; optical; Proteins; Research; Science; Computer software; Software; Spectrum Analysis; Spectroscopy; Spectrum Analyses; Technology; Temperature; Testing; Tissues; Body Tissues; Universities; Measures; Secondary Protein Structure; Label; Microscope; improved; Area; Phase; Biological; Biochemical; Spectroscopy, Fourier Transform Infrared; FTIR; FTIR spectroscopy; Chemicals; Chemical Structure; insight; Individual; Visible Radiation; Visible Light; Visible Light Radiation; Measurement; Plant Roots; root; Confocal Microscopy; infrared spectroscopy; instrument; Research Specimen; Specimen; Pulse; Physiologic pulse; anti-microbial susceptibility; Antimicrobial susceptibility; Dependence; Protocol; Protocols documentation; In Situ; Source; Techniques; Location; vibration; Degenerative Neurologic Diseases; Degenerative Neurologic Disorders; Nervous System Degenerative Diseases; Neural Degenerative Diseases; Neural degenerative Disorders; Neurodegenerative Diseases; Neurologic Degenerative Conditions; degenerative diseases of motor and sensory neurons; degenerative neurological diseases; neurodegenerative illness; Neurodegenerative Disorders; interest; infrared microscopy; Performance; fluorophore; Structure; novel; UV laboratory microscope; Ultraviolet Microscopes; fluorescence/UV microscope; fluorescent microscope; laboratory fluorescence light microscope; fluorescence microscope; Sampling; resistance to disease; resistant disease; resistant to disease; Disease Resistance; Address; Antimicrobial resistant; Resistance to antimicrobial; anti-microbial resistance; anti-microbial resistant; resistance to anti-microbial; resistant to anti-microbial; resistant to antimicrobial; Antimicrobial Resistance; Detection; multi-modal imaging; multi-modality imaging; multimodality imaging; Multimodal Imaging; Resolution; Optical Instrument; Optics/Optical Instrument; Small Business Innovation Research Grant; SBIR; Small Business Innovation Research; Preparation; Molecular; Development; developmental; ultraviolet; ultra violet; Image; imaging; National Institute of Biomedical Imaging and Bioengineering; NIBIB; protein misfolding; aberrant protein folding; abnormal protein folding; pathologic protein folding; protein mis-folding; optical imaging; optic imaging; anticancer research; anti-cancer research; cancer research; submicron; sub micron; spectroscopic imaging; imaging spectroscopy; antimicrobial; anti-microbial; biological research; fluorescence imaging; fluorescent imaging; prototype; multimodality; multi-modality; industry partner; industrial partnership; industry partnership; microscopic imaging; microscope imaging; microscopy imaging; feasibility testing

This Phase II project aims to develop and commercialize Fluorescence Enhanced Photothermal Infrared (FE-PTIR) imaging and spectroscopy. The proposed FE-PTIR will use fluorescence microscopy to map the distribution of fluorescently labeled regions of cells and tissue and then provide chemical structural analysis of the labeled regions using photothermal infrared spectroscopy. Fluorescence microscopy is a cornerstone technique in biological research, allowing sensitive mapping of specifically targeted biomolecules within cells and tissue, but it does not provide information about their molecular structure. Infrared (IR) spectroscopy can provide rich analysis of molecular structure and has been used in life sciences research to study tissue classification, drug/tissue interaction, neurodegenerative diseases, cancer and other areas. Conventional IR spectroscopy, however, has a fundamental spatial resolution limit (i.e. roughly how small an object it can analyze) of around micrometers, similar to the size of an average biological cell. Thus conventional IR spectroscopy has been extremely limited for many biomedical applications where the structures of interest are smaller than a cell. The FE-PTIR technique illuminates a fluorescently labeled sample with UV/visible light which results in fluorescent emission from fluorescently tagged molecules in the sample. A tunable infrared laser source also illuminates the sample, causing localized heating in the sample if the IR laser is tuned to a wavelength that excites molecular vibrations in the sample. Using a camera or other sensitive photodetector is used to record the fluorescent emission from different regions of the sample generates a map of the distribution of fluorescently labeled biomolecules. A key innovation of this proposal was the recognition that common fluorophores have an emission efficiency that is highly temperature dependent. Thus when the sample is also irradiated with infrared light at wavelengths corresponding to molecular vibrations, localized heating from IR absorption by the sample causes a significant change in the fluorescent emission. Recording the emission change as a function of sample position or IR wavelength produces IR absorption images and IR absorption spectra, respectively. Phase I research demonstrated the following key advances: (1) ability of FE-PTIR to map of target biomolecules with fluorescence and analyze the molecular structural of the target molecules; (2) achieve submicron spatial resolution for both fluorescence imaging and infrared spectroscopy; (3) demonstrated a 100X improvement in measurement sensitivity; (4) application of FE-PTIR to study of protein misfolding relevant to neurodegenerative disease research; (5) demonstrated FE-PTIR on individual bacterial and live cancer cells with subcellular resolution. This project is well aligned with NIH goals as it incorporates several key thrusts of the National Institute of Biomedical Imaging and Bioengineering, including optical imaging and spectroscopy, IR imaging, confocal microscopy, and multimodal imaging. FE- PTIR will be extremely useful for example in analyzing the molecular structure/folding/aggregation of fluorescence- localized proteins. Protein misfolding/aggregation is a root cause of many neurodegenerative diseases (e.g. Alzheimer's). Completion of this Phase II project will lead to the commercialization of a new multimodal microscope that will offer profound benefits for biomedical research including neurodegenerative diseases and antimicrobial resistance research.

Public Health Relevance Statement:
Project narrative Fluorescence microscopy is a cornerstone technique in biological research that maps the distribution of fluorescently labeled biomolecules but does not provide information on their chemical structure. Infrared spectroscopy provides detailed chemical structural analysis on biological materials, but fundamental resolution limits have constrained its application in biology. This SBIR project will overcome these limits to enable both mapping and chemical analysis of target biomolecules with sub-micron spatial resolution, thus enabling research to provide fundamental insights into neurodegenerative diseases and antimicrobial resistance.

Project Terms:
absorption; Alzheimer's Disease; AD dementia; Alzheimer; Alzheimer Type Dementia; Alzheimer disease; Alzheimer sclerosis; Alzheimer syndrome; Alzheimer's; Alzheimer's disease dementia; Alzheimers Dementia; Alzheimers disease; Primary Senile Degenerative Dementia; dementia of the Alzheimer type; primary degenerative dementia; senile dementia of the Alzheimer type; Amyloid; Amyloid Substance; Biocompatible Materials; Biomaterials; biological material; Biological Sciences; Biologic Sciences; Bioscience; Life Sciences; Biology; Biomedical Engineering; bio-engineered; bio-engineers; bioengineering; biological engineering; Biomedical Research; Malignant Neoplasms; Cancers; Malignant Tumor; malignancy; neoplasm/cancer; Cells; Cell Body; Classification; Systematics; Data Analyses; Data Analysis; data interpretation; Pharmacotherapy; Drug Therapy; drug treatment; Pharmaceutical Preparations; Drugs; Medication; Pharmaceutic Preparations; drug/agent; Fluorescence; Glucose; D-Glucose; Dextrose; Goals; Heating; Infrared Rays; infrared radiation; Institutes; Isotopes; Lasers; Laser Electromagnetic; Laser Radiation; Light; Photoradiation; Lipids; Maps; Metabolism; Intermediary Metabolism; Metabolic Processes; Fluorescence Microscopy; Fluorescence Light Microscopy; Molecular Structure; Macromolecular Structure; NIH; National Institutes of Health; United States National Institutes of Health; Nerve Cells; Nerve Unit; Neural Cell; Neurocyte; neuronal; Neurons; Proteins; Scientific Publication; Publications; Research; Software; Computer software; Spectroscopy; Spectrum Analyses; Spectrum Analysis; Technology; Temperature; Testing; Tissues; Body Tissues; Measures; tau Proteins; MT-bound tau; microtubule bound tau; microtubule-bound tau; tau; tau factor; τ Proteins; protein folding; Label; Microscope; improved; Area; Phase; Biological; biologic; Biochemical; Chemicals; Chemical Structure; insight; Individual; Visible Light; Visible Light Radiation; Visible Radiation; Measurement; root; Plant Roots; Confocal Microscopy; infrared spectroscopy; Malignant Cell; cancer cell; Research Specimen; Specimen; Pulse; Physiologic pulse; anti-microbial susceptibility; Antimicrobial susceptibility; Dependence; In Situ; Source; Techniques; brain tissue; vibration; Degenerative Neurologic Diseases; Degenerative Neurologic Disorders; Nervous System Degenerative Diseases; Neural Degenerative Diseases; Neural degenerative Disorders; Neurodegenerative Diseases; Neurologic Degenerative Conditions; degenerative diseases of motor and sensory neurons; degenerative neurological diseases; neurodegenerative illness; Neurodegenerative Disorders; interest; glucose metabolism; infrared microscopy; fluorophore; microbial; Structure; novel; Position; Positioning Attribute; Sampling; response; resistance to disease; resistant disease; resistant to disease; Disease Resistance; Photobleaching; Antimicrobial resistant; Resistance to antimicrobial; anti-microbial resistance; anti-microbial resistant; resistance to anti-microbial; resistant to anti-microbial; resistant to antimicrobial; Antimicrobial Resistance; Data; Detection; Molecular Analysis; multi-modal imaging; multi-modality imaging; multimodality imaging; Multimodal Imaging; Protein Analysis; Resolution; Optical Instrument; Optics/Optical Instrument; Small Business Innovation Research Grant; SBIR; Small Business Innovation Research; Molecular; Development; developmental; Image; imaging; protein misfolding; aberrant protein folding; abnormal protein folding; pathologic protein folding; optical imaging; optic imaging; design; designing; submicron; sub micron; spectroscopic imaging; imaging spectroscopy; innovation; innovate; innovative; antimicrobial; anti-microbial; data acquisition; biological research; fluorescence imaging; fluorescent imaging; engineering design; prototype; commercialization; multimodality; multi-modality; microscopic imaging; microscope imaging; microscopy imaging; biomedical imaging