Analysis of complex cellular processes in understanding disease, development of drugs, or testing efficacy of therapies often requires dynamic measurements of both cellular markers and cellular products. The ability to synchronously and rapidly conduct such measurements in complex cell- containing samples using a single instrumental analysis will be useful and beneficial across biotechnology. The goal of this project is to enable such multiplex analysis of T-cell activation using immunophenotyping and quantitative monitoring of their secreted cytokines as an important model system. High throughput measurement of samples of suspended cells is made possible using BennuBioâs Velocyt® multi-stream acoustic flow cytometry system. The primary innovation of this project is the implementation of negative acoustic contrast particles (NACPs) as a bead-based immunoassay platform that significantly extends the analytical capabilities of the Velocyt® instrument for simultaneous analysis of cell-associated and soluble biomarkers. In contrast to cells, which migrate to, and are analyzed at, discreet positions within the Velocyt® flow stream,NACPs are separated to distinct positions in the flow stream, allowing for easy discrimination between bead and cells in a single sample. In this proof-of-concept Phase I research, we aim to conduct simultaneous measurement of the T-cell activation marker CD25 in both the CD4 and CD8 population at the same time that we measure secreted interferon-γ and tumor necrosis factor-α . While this demonstration will be of significance to the development of pharmaceuticals for treatment of a range of immunological and infectious diseases that involve T-cell activation, it will also provide a firm basis by which to design and commercialize customizable NACP-based reagent kits to address a wide range of disciplines across biotechnology.
Public Health Relevance Statement: Project Narrative This project will establish the feasibility of developing a new bead-based reagent technology for use with BennuBioâs Velocyt® high throughput acoustic flow cytometry system. This technology will augment the Velocytâs® rapid cell analysis capabilities to enable simultaneous quantitative measurement of cell-secreted biomarkers. Concurrent, multiplex analysis of cell-borne and secreted, soluble biomarkers will simplify the development of therapeutics targeting a wide range of diseases and enable important new capabilities across biotechnology.
Project Terms: Acoustic; Acoustics; Antibodies; Autoimmune Diseases; autoimmune condition; autoimmune disorder; autoimmunity disease; Biological Assay; Assay; Bioassay; Biologic Assays; Biomedical Research; Biotechnology; Biotech; Biotin; Vitamin H; coenzyme R; Blood; Blood Reticuloendothelial System; Blood Cells; Peripheral Blood Cell; Cell physiology; Cell Function; Cell Process; Cellular Function; Cellular Physiology; Cellular Process; Subcellular Process; Cells; Cell Body; Communicable Diseases; Infectious Disease Pathway; Infectious Diseases; Infectious Disorder; Diagnosis; Discrimination; Cognitive Discrimination; Disease; Disorder; Elastomers; Engineering; Exhibits; Flow Cytometry; Flow Cytofluorometries; Flow Cytofluorometry; Flow Microfluorimetry; Flow Microfluorometry; flow cytophotometry; Fluorescence; Goals; Human; Modern Man; Immobilization; orthopedic freezing; Immunoassay; Immune System Diseases; Immune Diseases; Immune Disorders; Immune Dysfunction; Immune System Disorder; Immune System Dysfunction; Immune System and Related Disorders; Immunodeficiency and Immunosuppression Disorders; Immunologic Diseases; Immunological Diseases; Immunological Dysfunction; Immunological System Dysfunction; Interferon Type II; IFN-Gamma; IFN-g; IFN-γ; IFNG; IFNγ; Immune Interferon; Interferon Gamma; lFN-Gamma; Interferons; IFN; Lasers; Laser Electromagnetic; Laser Radiation; Methods; Methodology; Microspheres; Microbeads; Biological Models; Biologic Models; Model System; Plasma; Blood Plasma; Plasma Serum; Reticuloendothelial System, Serum, Plasma; pressure; Proteins; Reagent; Research; Silicones; Specificity; Suspensions; Suspension substance; T-Lymphocyte; T-Cells; thymus derived lymphocyte; Technology; Testing; Time; tissue culture; Travel; Up-Regulation; Upregulation; Work Simplification; Streptavidin; Strepavidin; cytokine; Immunophenotyping; Immunologic Subtyping; immunophenotype; Measures; CD3; CD3 Complex; CD3 molecule; OKT3 antigen; T3 Antigens; T3 Complex; T3 molecule; CD3 Antigens; Electrical Impedance; Impedance; electric impedance; Surface; Clinical; Microscopic; Phase; Variation; Variant; biologic; Biological; Blood Serum; Serum; insight; Discipline; Measurement; CD8 Cell; CD8 T cells; CD8 lymphocyte; CD8+ T cell; CD8+ T-Lymphocyte; CD8-Positive Lymphocytes; T8 Cells; T8 Lymphocytes; CD8-Positive T-Lymphocytes; Oncology Cancer; Oncology; fluid; liquid; Liquid substance; instrument; Frequencies; Complex; Stream; elastomeric; System; Width; particle; Performance; receptor; Receptor Protein; solute; Peripheral Blood Mononuclear Cell; PBMC; aqueous; Speed; novel; Cell surface; Positioning Attribute; Position; TNF gene; (TNF)-α; Cachectin; Macrophage-Derived TNF; Monocyte-Derived TNF; TNF; TNF A; TNF Alpha; TNF-α; TNFA; TNFα; Tumor Necrosis Factor; Tumor Necrosis Factor-alpha; Cytometry; Modeling; Sampling; Property; drug development; drug discovery; activate T cells; T-Cell Activation; Pharmaceutical Agent; Pharmaceuticals; Pharmacological Substance; pharmaceutical; Pharmacologic Substance; Cell secretion; Cellular Secretion; CD28 gene; CD28; T44; IL2RA gene; CD25; IL2R; IL2RA; TCGFR; CD8B1 gene; CD8; CD8B; CD8B1; LYT3; Address; Data; Detection; Protein Analysis; Supporting Cell; Monitor; Preparation; preparations; Process; Development; developmental; Pathway interactions; pathway; designing; design; intervention efficacy; therapeutic efficacy; therapy efficacy; Treatment Efficacy; Population; migration; Coupled; innovate; innovative; innovation; Cellular model; Cell model; therapeutic target; commercialization; stem cell technology; therapeutic agent development; therapeutic development; bio-markers; biologic marker; biomarker; Biological Markers; efficacy testing; specific biomarkers; protein markers; protein biomarkers; detection procedure; detection technique; detection method; Prognosis; multiplex assay; diagnostic value; diagnostic ability; diagnostic capability; diagnostic power; d
