Confocal imaging of slit diaphragm proteins in expanded kidney tissue

• The subtlest element of the kidney, such as the slit diaphragm, has historically only been spatially resolved using electron microscopy due to the nanometer-scale dimensions of these structures. Recently, it was shown that the nanoscale distribution of proteins in the slit diaphragm can be resolved by fluorescence based stimulated emission depletion (STED) microscopy, in combination with optical clearing. Fluorescence microscopy has advantages over electron microscopy in terms of multiplex imaging of different molecular species (i.e. epitopes), and also the amount of volumetric data that can be extracted from a thick sample. However, STED microscopy is still a costly technique commonly not available to all life science researchers. An image technique with which the slit diaphragm proteins in the kidney can be visualized using more standard fluorescence imaging techniques is thus desirable. Recent studies have shown that biological tissue samples can be isotropically expanded while optically cleared, revealing nanoscale localizations of multiple epitopes using confocal microscopy. Here we show that kidney samples can be expanded sufficiently to study the finest elements of the filtration barrier under both healthy and diseased conditions using confocal microscopy. This finding opens up the possibility for any researcher with access to a confocal microscope to study foot process protein distributions on the effective nanometer-scale. We also show that expansion microscopy can be combined with STED microscopy to further increase the effective spatial resolution down to below 20 nm.

Ämnesord

NATURVETENSKAP  -- Biologi -- Biofysik (hsv//swe)

NATURAL SCIENCES  -- Biological Sciences -- Biophysics (hsv//eng)

Nyckelord

Expansion microscopy

kidney

slit diaphragm

Super-resolution microscopy

Biological Physics

Biologisk fysik

Publikations- och innehållstyp

vet (ämneskategori)

ovr (ämneskategori)