Issue 45, 2023

Differential labelling of human sub-cellular compartments with fluorescent dye esters and expansion microscopy

Abstract

Amine-reactive esters of aromatic fluorescent dyes are emerging as imaging probes for nondescript staining of cellular and tissue architectures. We characterised the staining patterns of 14 fluorescent dye ester species with varying physical and spectral properties in the broadly studied human HeLa cell line. When combined with the super-resolution technique expansion microscopy (ExM) involving swellable acrylamide hydrogels, fluorescent esters reveal nanoscale features including cytoplasmic membrane-bound compartments and nucleolar densities. We observe differential labelling patterns linked to the biochemical properties of the conjugated dye. Alterations in staining density and compartment specificity were seen depending on the timepoint of application in the ExM protocol. Additional complexity in labelling patterns was detected arising from inter-ester interactions. Our findings raise a number of considerations for the use of fluorescent esters. We demonstrate esters as a useful addition to the repertoire of stains of the cellular proteome, whether applied either on their own to visualise overall cellular morphology, or as counterstains providing ultrastructural context alongside specific target markers like antibodies.

Graphical abstract: Differential labelling of human sub-cellular compartments with fluorescent dye esters and expansion microscopy

Supplementary files

Article information

Article type
Paper
Submitted
10 mar 2023
Accepted
07 okt 2023
First published
09 noy 2023
This article is Open Access
Creative Commons BY license

Nanoscale, 2023,15, 18489-18499

Differential labelling of human sub-cellular compartments with fluorescent dye esters and expansion microscopy

T. M. D. Sheard, T. B. Shakespeare, R. S. Seehra, M. E. Spencer, K. M. Suen and I. Jayasinghe, Nanoscale, 2023, 15, 18489 DOI: 10.1039/D3NR01129A

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