The immunological synapse (IS) is a specialised cell-cell adhesion that mediates antigen acquisition and regulates the activation of lymphocytes. Initial studies of the IS showed a structure composed of stable supra-molecular activation clusters (SMAC) organised during the interaction of helper T lymphocytes with B lymphocytes, working as antigen presenting cells. A central SMAC of coalesced T cell receptors (TCRs) and a peripheral SMAC for cell-cell adhesion were observed. IS with similar structure was later described during antigen acquisition by B cells and during the interaction of NK cells with target and healthy cells. More recent research developed with microscopy systems that improve the spatial and temporal resolution has showed the complex molecular dynamics at the IS that governs lymphocyte activation. Currently, the IS is seen as a three-dimensional structure where signalling networks for lymphocyte activation and endosomal and cytoskeleton machinery are polarised. A view has emerged in which dynamic microclusters of signalling complexes are composed of molecular components attached to the plasma membrane and other components conveyed on sub-synaptic vesicles transported to the membrane by cytoskeletal fibers and motor proteins. Much information is nonetheless missing about how the dynamics of the endosomal compartment, the cytoskeleton, and signalling complexes are reciprocally regulated to achieve the function of lymphocytes. Experimental evidence also suggests that the environment surrounding lymphocytes exposed to different antigenic challenge regulates IS assembly and functional output, making an even more complex scenario still far from being completely understood. Also, although some signalling molecular components for lymphocyte activation have been identified and thoroughly studied, the function of other molecules has not been yet uncovered or deeply characterised. This research topic aims to provide the reader with the latest information about the molecular dynamics governing lymphocyte activation. These molecular dynamics dictate cell decisions. Thus, we expect that understanding them will provide new avenues for cell manipulation in therapies to treat different immune-related pathologies.

The immunological synapse (IS) is a specialised cell-cell adhesion that mediates antigen acquisition and regulates the activation of lymphocytes. Initial studies of the IS showed a structure composed of stable supra-molecular activation clusters (SMAC) organised during the interaction of helper T lymphocytes with B lymphocytes, working as antigen presenting cells. A central SMAC of coalesced T cell receptors (TCRs) and a peripheral SMAC for cell-cell adhesion were observed. IS with similar structure was later described during antigen acquisition by B cells and during the interaction of NK cells with target and healthy cells. More recent research developed with microscopy systems that improve the spatial and temporal resolution has showed the complex molecular dynamics at the IS that governs lymphocyte activation. Currently, the IS is seen as a three-dimensional structure where signalling networks for lymphocyte activation and endosomal and cytoskeleton machinery are polarised. A view has emerged in which dynamic microclusters of signalling complexes are composed of molecular components attached to the plasma membrane and other components conveyed on sub-synaptic vesicles transported to the membrane by cytoskeletal fibers and motor proteins. Much information is nonetheless missing about how the dynamics of the endosomal compartment, the cytoskeleton, and signalling complexes are reciprocally regulated to achieve the function of lymphocytes. Experimental evidence also suggests that the environment surrounding lymphocytes exposed to different antigenic challenge regulates IS assembly and functional output, making an even more complex scenario still far from being completely understood. Also, although some signalling molecular components for lymphocyte activation have been identified and thoroughly studied, the function of other molecules has not been yet uncovered or deeply characterised. This research topic aims to provide the reader with the latest information about the molecular dynamics governing lymphocyte activation. These molecular dynamics dictate cell decisions. Thus, we expect that understanding them will provide new avenues for cell manipulation in therapies to treat different immune-related pathologies.

This new collection features the most up-to-date essential protocols that are currently being used to study the immune synapse. Beginning with methods for making biophysical measurements, the volume continues by covering the cell biology of synapses, methods for advanced substrate engineering, mechanobiology topics, new technologies to describe and manipulate synaptic components, as well as methods related to sites of action and immunotherapy. Written for the highly successful Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step and readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and fully updated, The Immune Synapse: Methods and Protocols, Second Edition serves as an ideal practical guide for researchers working in this dynamic field. Chapters 5, 11,18, 27, 30, and 32 are available open access under a Creative Commons Attribution 4.0 International License via link.springer.com.

This new collection features the most up-to-date essential protocols that are currently being used to study the immune synapse. Beginning with methods for making biophysical measurements, the volume continues by covering the cell biology of synapses, methods for advanced substrate engineering, mechanobiology topics, new technologies to describe and manipulate synaptic components, as well as methods related to sites of action and immunotherapy. Written for the highly successful Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step and readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and fully updated, The Immune Synapse: Methods and Protocols, Second Edition serves as an ideal practical guide for researchers working in this dynamic field. Chapters 5, 11, 18, 27, 30, and 32 are available open access under a Creative Commons Attribution 4.0 International License via link.springer.com.

Finally, vesicle cargo was shown to correlate with different vesicle populations, based on their membrane order, demonstrating vesicle order may mirror the heterogeneous nature of the PM. These results demonstrate the complex biophysical processes that control the T cell immunological synapse are an amalgamation of cytoskeletal organisation, vesicle dynamics and membrane polarity.

The Immunological Synapse, Part A, Volume 173 in the Methods in Cell Biology series provides state-of-the-art methods for the study of the immunological synapse. Sections cover Imaging polarized granule release at the cytotoxic T cell immunological synapse using TIRF microscopy: control by polarity regulators, Analysis of centrosomal area actin reorganization and centrosome polarization upon lymphocyte activation at the immunological synapse, P815-based redirected degranulation assay to study human NK cell effector functions, Cytotoxic and Chemotactic Dynamics of Natural Killer Cells Quantified by Live-cell Imaging, Quantification of interaction frequency between antigen-presenting cells and T cells by conjugation assay, and more. Other chapters focus on the Study of the Effects of NK-Tumor Cell Interaction by Proteomic Analysis and Imaging, Quantification of lymphocytic choriomeningitis virus specific T cells and LCMV viral titers, Quantification of lymphocytic choriomeningitis virus specific T cells and LCMV viral titers, An in vitro model to monitor natural killer cell effector functions against primary breast cancer, and Standardized Protocol for the Evaluation of Chimeric Antigen Receptor (CAR)-modified Cell Immunological Synapse Quality using the Glass-supported Planar L. - Covers various methods related to the study of the immunological synapse - Includes detailed, point-by-point, methods as well as various important notes - Provides the authority and expertise from an international board of leading scientists