Elispot for Rookies (and Experts Too) - Janetzki, Sylvia; SpringerLink (Online service) (Cham : Springer International Publishing : Imprint: Springer, 2016)
Preface -- Overview -- Historic Overview -- Principles of Elispot -- Modifications -- Simplifications -- Coating Step -- Detection Step -- Adaptations -- Detection of specific antibodies -- Detection of multiple analytes in one well -- Dual color Elispot -- Fluorospot -- Summary -- Important Assay Details -- Reagents and Materials -- Elispot plates -- Antibody choices -- Enzymatic conjugates and substrates -- Fluorescent dyes -- Culture media -- Blocking buffers -- Washing buffers -- Important laboratory supplies -- Pipettes -- Filters -- Other supplies -- Automated Elispot reader -- Sample preparation -- Overview -- Whole blood and PBMC -- Blood draw and anticoagulants -- PBMC isolation -- Time frame between blood draw and PBMC isolation -- Immune cells obtained from tissue -- Freezing of cells -- Thawing of cells -- Cell counting and viability -- Overnight resting -- Isolation of sub-populations of cells -- Expansion of cells -- Planning the experiment -- Controls -- Negative controls -- Positive controls -- External controls -- Plate layout -- Timing -- The Elispot assay -- Coating -- Blocking -- Effector cell preparation -- Stimulant preparation -- Peptides and peptide pools -- Proteins and lysate -- Antigen presenters -- Control stimulants -- Reference samples -- Plating out cells and stimulants -- Incubation -- Removal of cells -- Spot development -- Enumeration of spots -- Data analysis -- Quality control -- Reporting guidelines -- Troubleshooting -- References.
Keywords: Immunology; Cell biology; Laboratory medicine; Immunology; Cell Biology; Laboratory Medicine