Super-resolution imaging of the natural killer cell immunological synapse on a glass-supported planar lipid bilayer.

Journal of Visualized Experiments : JoVE
Peilin ZhengDongfang Liu

Abstract

The glass-supported planar lipid bilayer system has been utilized in a variety of disciplines. One of the most useful applications of this technique has been in the study of immunological synapse formation, due to the ability of the glass-supported planar lipid bilayers to mimic the surface of a target cell while forming a horizontal interface. The recent advances in super-resolution imaging have further allowed scientists to better view the fine details of synapse structure. In this study, one of these advanced techniques, stimulated emission depletion (STED), is utilized to study the structure of natural killer (NK) cell synapses on the supported lipid bilayer. Provided herein is an easy-to-follow protocol detailing: how to prepare raw synthetic phospholipids for use in synthesizing glass-supported bilayers; how to determine how densely protein of a given concentration occupies the bilayer's attachment sites; how to construct a supported lipid bilayer containing antibodies against NK cell activating receptor CD16; and finally, how to image human NK cells on this bilayer using STED super-resolution microscopy, with a focus on distribution of perforin positive lytic granules and filamentous actin at NK synapses. Thus, combining...Continue Reading

Citations

Sep 14, 2018·Journal of Cell Science·Edward JenkinsErdinc Sezgin
Feb 6, 2019·International Journal of Molecular Sciences·Guillaume Le Saux, Mark Schvartzman
Jul 19, 2020·The FEBS Journal·Pablo F CéspedesErdinc Sezgin
Sep 4, 2015·Current Protocols in Cell Biology·Travis J CritesRajat Varma
Aug 28, 2020·Cell Communication and Signaling : CCS·Dongfang LiuChen Liu

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Methods Mentioned

BETA
imaging techniques
electron microscopy
FACS
fluorescence
confocal microscopy
flow cytometry

Software Mentioned

FACS analysis
Huygens

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