XIV, 322 p. 123 illus., 10 illus. in color. online resource
SUMMARY
Although the ELISPOT (enzyme-linked immunospot) assay is widely used in many fields of biomedical research, producing recognizable and quantifiable spots may be very difficult not only for beginners, but also for experienced researchers. In Handbook of ELISPOT: Methods and Protocols, the first book dedicated entirely to the ELISPOT, recognized experts with first-hand experience detail how to design, perform, and analyze these assays. The readily reproducible techniques they provide cover a wide variety of topics, including the use of membrane-backed plates, the standardization and validation procedures, the removal of cells from ELISPOT plates, cell separation techniques, and the quantification of ELISPOT data. There are also numerous ELISPOT applications involving animal models, human cells, measles, multiple sclerosis, immune responses, multicytokine detection systems, and immunocytochemistry. Highlights include dual-color and multiplex ELISPOT assays, use of the ELISPOT assay on feline lymphocytes, standardization of the ELISPOT procedure, and combining the ELISPOT assay with immunohistochemistry. The protocols follow the successful Methods in Molecular Biology™ series format, each offering step-by-step laboratory instructions, an introduction outlining the principle behind the technique, lists of the necessary equipment and reagents, and tips on troubleshooting and avoiding known pitfalls. Comprehensive and highly practical, Handbook of ELISPOT: Methods and Protocols offers both novice and experienced researchers a wealth of easy-to-understand information that will clarify the technically challenging aspects of ELISPOT and open the door to its wider successful use
CONTENT
Elispot Assay Setup and Data Analysis -- ELISPOT Assay -- Chemistry and Biology of the ELISPOT Assay -- Membranes and Membrane Plates Used in ELISPOT -- Standardization and Validation Issues of the ELISPOT Assay -- A Cell-Detachment Solution Can Reduce Background Staining in the ELISPOT Assay -- Isolation of Subsets of Immune Cells -- Image Analysis and Data Management of ELISPOT Assay Results -- High Resolution as a Key Feature to Perform Accurate ELISPOT Measurements Using Zeiss KS ELISPOT Readers -- Elispot Assay in Animal Models -- Improving the Sensitivity of the ELISPOT Analyses of Antigen-Specific Cellular Immune Responses in Rhesus Macaques -- Feline Cytokine ELISPOT -- Measuring T-Cell Function in Animal Models of Tuberculosis by ELISPOT -- Interferon-? ELISPOT Assay for the Quantitative Measurement of Antigen-Specific Murine CD8+ T-Cells -- Elispot Assays in Human Studies -- Detection of Measles Virus-Specific Interferon-?-Secreting T-Cells by ELISPOT -- Epitope Mapping in Multiple Sclerosis Using the ELISPOT Assay -- Use of Interferon-? ELISPOT in Monitoring Immune Responses in Humans -- ELISPOT Determination of Interferon-? T-Cell Frequencies in Patients With Autoimmune Sensorineural Hearing Loss -- Multiplex and Modified Elispot Assay Formats -- Dual-Color ELISPOT Assay for Analyzing Cytokine Balance -- Simultaneous Detection of Multiple Cytokines in ELISPOT Assays -- Fluorospot Assay -- A Gel-Based Dual Antibody Capture and Detection Method for Assaying of Extracellular Cytokine Secretion
