Skip to Content
Merck
  • Subdiffraction-resolution fluorescence imaging of immunological synapse formation between NK cells and A. fumigatus by expansion microscopy.

Subdiffraction-resolution fluorescence imaging of immunological synapse formation between NK cells and A. fumigatus by expansion microscopy.

Communications biology (2021-10-06)
Nora Trinks, Sebastian Reinhard, Matthias Drobny, Linda Heilig, Jürgen Löffler, Markus Sauer, Ulrich Terpitz
ABSTRACT

Expansion microscopy (ExM) enables super-resolution fluorescence imaging on standard microscopes by physical expansion of the sample. However, the investigation of interactions between different organisms such as mammalian and fungal cells by ExM remains challenging because different cell types require different expansion protocols to ensure identical, ideally isotropic expansion of both partners. Here, we introduce an ExM method that enables super-resolved visualization of the interaction between NK cells and Aspergillus fumigatus hyphae. 4-fold expansion in combination with confocal fluorescence imaging allows us to resolve details of cytoskeleton rearrangement as well as NK cells' lytic granules triggered by contact with an RFP-expressing A. fumigatus strain. In particular, subdiffraction-resolution images show polarized degranulation upon contact formation and the presence of LAMP1 surrounding perforin at the NK cell-surface post degranulation. Our data demonstrate that optimized ExM protocols enable the investigation of immunological synapse formation between two different species with so far unmatched spatial resolution.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Sodium acrylate, 97%
Sigma-Aldrich
Sodium chloride, BioReagent, suitable for cell culture, suitable for insect cell culture, suitable for plant cell culture, ≥99%
Sigma-Aldrich
Anti-Mouse IgG (H+L), highly cross adsorbed antibody produced in goat, affinity isolated antibody, buffered aqueous solution
Sigma-Aldrich
Histopaque®-1077, sterile-filtered, density: 1.077 g/mL
Sigma-Aldrich
Formaldehyde solution, for molecular biology, 36.5-38% in H2O
Sigma-Aldrich
Guanidine hydrochloride, BioUltra, for molecular biology, ≥99.5% (AT)
Sigma-Aldrich
N,N,N′,N′-Tetramethylethylenediamine, BioReagent, for molecular biology, ≥99% (GC)
Sigma-Aldrich
Ammonium persulfate, for molecular biology, suitable for electrophoresis, ≥98%