Issue 7, 2019

Rhodamine–Hoechst positional isomers for highly efficient staining of heterochromatin

Abstract

Hoechst conjugates to fluorescent dyes are popular DNA stains for live-cell imaging, but the relationship between their structure and performance remains elusive. This study of carboxyrhodamine–Hoechst 33258 conjugates reveals that a minimal change in the attachment point of the dye has dramatic effects on the properties of the final probe. All tested 6′-carboxyl dye-containing probes exhibited dual-mode binding to DNA and formed a dimmer complex at high DNA concentrations. The 5′-carboxyl dye-containing probes exhibited single-mode binding to DNA which translated into increased brightness and lower cytotoxicity. Up to 10-fold brighter nuclear staining by the newly developed probes allowed acquisition of stimulated emission depletion (STED) nanoscopy images of outstanding quality in living and fixed cells. Therefore we were able to estimate a diameter of ∼155 nm of the heterochromatin exclusion zones in the nuclear pore region in living cells and intact chicken erythrocytes and to localize telomeres relative to heterochromatin in living U-2 OS cells. Employing the highly efficient probes for two-color STED allowed visualization of DNA and tubulin structures in intact nucleated erythrocytes – a system where imaging is greatly hampered by high haemoglobin absorbance.

Graphical abstract: Rhodamine–Hoechst positional isomers for highly efficient staining of heterochromatin

Supplementary files

Article information

Article type
Edge Article
Submitted
14 Nov 2018
Accepted
11 Dec 2018
First published
12 Dec 2018
This article is Open Access

All publication charges for this article have been paid for by the Royal Society of Chemistry
Creative Commons BY license

Chem. Sci., 2019,10, 1962-1970

Rhodamine–Hoechst positional isomers for highly efficient staining of heterochromatin

J. Bucevičius, J. Keller-Findeisen, T. Gilat, S. W. Hell and G. Lukinavičius, Chem. Sci., 2019, 10, 1962 DOI: 10.1039/C8SC05082A

This article is licensed under a Creative Commons Attribution 3.0 Unported Licence. You can use material from this article in other publications without requesting further permissions from the RSC, provided that the correct acknowledgement is given.

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