Polychromatic flow cytometry [FC] is one of the most powerful analytical techniques routinely used in immunology. Flow cytometry plays a critical role in several growing clinical applications: leukemia and lymphoma immunophenotyping for diagnosis and sub-categorization (now using 8-10 colors);in CD4/CD8 measurement in HIV-positive patients;and in HLA typing for transplantation cross-matching. With the increasing incidence of hematologic malignancies and use of transplantation, the number of FC tests will continue to grow as reliance on the methodology deepens. Increasing the number of fluorophores on a single sample will give clinicians a higher level of accuracy in the diagnosis and management of certain pediatric hematological disorders, where sample size is limited, and even allow for sub-categorization of the leukemia or lymphoma. Further utility of additional fluorophores for these polychromatic panels is for diagnostic samples of small volume such as specimens from an ever-increasing number of minimally invasive procedures that produce limited volume specimens, such as fine needle aspiration, laparoscopy, core biopsy, endoscopy, bronchoscopy, and notoriously hypocellular cerebrospinal fluid. Due to the small sample size of most diagnostic specimens in current practice, often insufficient cell numbers are available for the necessarily numerous panels to reach a diagnosis. A uniform deficiency of most common fluorophores is their broad fluorescence [emission bands], which cause spectral overlap, requiring extensive pre-assay experimentation and use of mathematical compensation, all of which increase experimental error, reduce sensitivity, and limit multiplexing. NIRvana Sciences, Inc. proposes to develop the first three members of a palette of water-soluble, conjugatable, bright, emission wavelength-tunable bacteriochlorin fluorophores characterized by sharp emission bands in the near-infrared [NIR] region upon excitation by commonly used laser wavelengths. The new fluorophores are based on a platform technology enabled by concise, versatile synthetic routes to bacteriochlorins with the following features: photo and thermal stability;rational control of excitation and emission band positions;additional substitution for water solubility and bioconjugation;and light absorption at laser lines of 355, 532, and 561 nm [with corresponding intense NIR emission bands]. The bacteriochlorins exhibit long """"""""Stokes'-like shifts"""""""" as all three absorption bands produce the same strong emission in the NIR region. These new fluorescent dyes fill an important unmet need in cancer diagnosis as well as pediatric and other limited-sample applications.

Public Health Relevance

Flow Cytometry is used routinely for both basic research and clinical diagnostic applications. Human blood cancers, many types of infectious diseases, and autoimmunity are routinely diagnosed using Flow Cytometry. The advent of newer sharper dyes will allow for higher sensitivity, smaller sample size, higher throughput testing, and reduced costs for the diagnosis of these diseases.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Small Business Technology Transfer (STTR) Grants - Phase I (R41)
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Special Emphasis Panel (ZRG1-IMST-K (14))
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Prograis, Lawrence J
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Nirvana Sciences, Inc.
Research Triangle Park
United States
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