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Simultaneous multi-lifetime multi-color STED imaging for colocalization analyses.

Optics express (2011-03-04)
Johanna Bückers, Dominik Wildanger, Giuseppe Vicidomini, Lars Kastrup, Stefan W Hell
ABSTRACT

We describe a STED microscope optimized for colocalization experiments with up to three colors. Two fluorescence labels are separated by their fluorescence lifetime whereas a third channel is discriminated by the wavelength of fluorescence emission. Since it does not require a second STED beam, separating by lifetime is insensitive to drift and thus optimally suited for colocalization analyses. Furthermore, we propose a setup having a second STED beam for long duration multicolor recording.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
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Atto 647N azide
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Phalloidin–Atto 532, suitable for fluorescence, ≥95.0% (HPCE)
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Atto 532 maleimide, BioReagent, suitable for fluorescence, ≥90% (coupling to thiols)
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Atto 532 DOPE, suitable for fluorescence
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Atto 425 maleimide, BioReagent, suitable for fluorescence, ≥70% (coupling to thiols)
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Atto 425 azide, suitable for fluorescence

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