Handbook of ELISPOT

In this first book dedicated entirely to the ELISPOT, a critical enzyme-linked immunospot assay used widely in biomedical research, 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.

Part I. ELISPOT Assay Setup and Data Analysis The ELISPOT Assay: A Personal Retrospective Jonathon D. Sedgwick Chemistry and Biology of the ELISPOT Assay Alexander E. Kalyuzhny Membranes and Membrane Plates Used in ELISPOT Alan J. Weiss Standardization and Validation Issues of the ELISPOT Assay Sylvia Janetzki, Josephine H. Cox, Neal Oden, and Guido Ferrari A Cell-Detachment Solution Can Reduce Background Staining in the ELISPOT Assay Angela Grant, Sarah Palzer, Chris Hartnett, Tanya Bailey, Monica Tsang, and Alexander E. Kalyuzhny Isolation of Subsets of Immune Cells Carrie E. Peters, Steven M. Woodside, and Allen C. Eaves Image Analysis and Data Management of ELISPOT Assay Results Paul Viktor Lehmann High Resolution as a Key Feature to Perform Accurate ELISPOT Measurements Using Zeiss KS ELISPOT Readers Wolf Malkusch Part II. ELISPOT Assay in Animal Models Improving the Sensitivity of the ELISPOT Analyses of Antigen-Specific Cellular Immune Responses in Rhesus Macaques K. Jagannadha Sastry, Pramod N. Nehete, and Bharti Nehete Feline Cytokine ELISPOT: Issues in Assay Development Sushila K. Nordone, Rosemary Stevens, Alora S. LaVoy, and Gregg A. Dean Measuring T-Cell Function in Animal Models of Tuberculosis by ELISPOT Vanja Lazarevic, Santosh Pawar, and JoAnne Flynn Interferon-g ELISPOT Assay for the Quantitative Measurement of Antigen-Specific Murine CD8+ T-Cells Geoffrey A. Cole Part III. ELISPOT Assays in Human Studies Detection of Measles Virus-Specific Interferon-g-Secreting T-Cells by ELISPOT Jenna E. Ryan, Inna G. Ovsyannikova, and Gregory A. Poland Epitope Mapping in Multiple Sclerosis Using the ELISPOT Assay Clara M. Pelfrey and Ioana R. Moldovan Use of Interferon-g ELISPOT in Monitoring Immune Responsesin Humans Mark Matijevic and Robert G. Urban ELISPOT Determination of Interferon-g T-Cell Frequencies in Patients With Autoimmune Sensorineural Hearing Loss C. Arturo Solares and Vincent K. Tuohy Part IV. Multiplex and Modified ELISPOT Assay Formats Dual-Color ELISPOT Assay for Analyzing Cytokine Balance Yoshihiro Okamoto and Mikio Nishida Simultaneous Detection of Multiple Cytokines in ELISPOT Assays Sarah Palzer, Tanya Bailey, Chris Hartnett, Angela Grant, Monica Tsang, and Alexander E. Kalyuzhny Fluorospot Assay: Methodological Analysis Agnes Gazagne, Wolf Malkusch, Benoit Vingert, Wolf H. Fridman, and Eric Tartour A Gel-Based Dual Antibody Capture and Detection Method for Assaying of Extracellular Cytokine Secretion: EliCell Lisa A. Spencer, Rossana C. N. Melo, Sandra A. C. Perez, and Peter F. Weller Index