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Issue 7, 2019
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Rhodamine–Hoechst positional isomers for highly efficient staining of heterochromatin

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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

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Publication details

The article was received on 14 Nov 2018, accepted on 11 Dec 2018 and first published on 12 Dec 2018


Article type: Edge Article
DOI: 10.1039/C8SC05082A
Chem. Sci., 2019,10, 1962-1970
  • Open access: Creative Commons BY license
    All publication charges for this article have been paid for by the Royal Society of Chemistry

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    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

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