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Imaging the Human Immunological Synapse
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Imaging the Human Immunological Synapse.

Ana Bello-Gamboa1, Juan Manuel Izquierdo1, Marta Velasco1

  • 1Instituto de Investigaciones Biomédicas Alberto Sols, CSIC-Universidad Autónoma de Madrid; Departamento de Bioquímica, Instituto de Investigaciones Biomédicas Alberto Sols CSIC-UAM, Facultad de Medicina, Universidad Autónoma de Madrid.

Journal of Visualized Experiments : Jove
|January 14, 2020
PubMed
Summary

This study details a method for generating and imaging immunological synapses (IS) between antigen-presenting cells (APCs) and T cells. The technique enhances image quality and temporal resolution for studying cell-cell interactions.

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Area of Science:

  • Immunology
  • Cell Biology
  • Microscopy

Background:

  • Immunological synapses (IS) are crucial for adaptive immune responses, mediating communication between antigen-presenting cells (APCs) and T lymphocytes.
  • Understanding the dynamic events during IS formation is essential for deciphering immune cell function and dysfunction.
  • Current imaging techniques face challenges in capturing the early stages and complex dynamics of IS formation in a biologically relevant context.

Purpose of the Study:

  • To develop and validate a robust protocol for generating and imaging immunological synapses (IS).
  • To capture and analyze the initial formation stages and subsequent trafficking events within APC-T cell conjugates.
  • To provide a method that enhances image quality and temporal resolution for studying IS dynamics.

Main Methods:

  • Utilized Jurkat cells and Staphylococcus enterotoxin E (SEE)-pulsed Raji cells to model T cell-APC synaptic conjugates.
  • Employed cell-to-cell conjugation followed by time-lapse acquisition using wide-field fluorescence microscopy (WFFM).
  • Applied post-acquisition deconvolution for image processing to improve signal-to-noise ratio and temporal resolution.

Main Results:

  • Achieved enhanced signal-to-noise ratio and temporal resolution in IS imaging.
  • Enabled synchronized acquisition of multiple fluorochromes in forming synaptic conjugates.
  • Demonstrated compatibility with downstream immunofluorescence staining and advanced microscopy techniques like LSCM.

Conclusions:

  • The presented protocol effectively models and visualizes key events in immunological synapse formation.
  • The method offers improved imaging capabilities for studying complex cell-cell interactions in a near-physiological setting.
  • This approach provides a valuable tool for immunological research, despite limitations in Z-axis imaging.