Nonlinear
Emission of Quinolizinium-Based Dyes with
Application in Fluorescence Lifetime Imaging
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Abstract
Charged
molecules based on the quinolizinum cation have potential
applications as labels in fluorescence imaging in biological media
under nonlinear excitation. A systematic study of the linear and nonlinear
photophysics of derivatives of the quinolizinum cation substituted
by either dimethylaniline or methoxyphenyl electron donors is performed.
The effects of donor strength, conjugation length, and symmetry in
the two-photon emission efficiency are analyzed in detail. The best
performing nonlinear fluorophore, with two-photon absorption cross
sections of 1140 GM and an emission quantum yield of 0.22, is characterized
by a symmetric D-π-A<sup>+</sup>-π-D architecture based
on the methoxyphenyl substituent. Application of this molecule as
a fluorescent marker in optical microscopy of living cells revealed
that, under favorable conditions, the fluorophore can be localized
in the cytoplasmatic compartment of the cell, staining vesicular shape
organelles. At higher dye concentrations and longer staining times,
the fluorophore can also penetrate into the nucleus. The nonlinearly
excited fluorescence lifetime imaging shows that the fluorophore lifetime
is sensitive to its location in the different cell compartments. Using
fluorescence lifetime microscopy, a multicolor map of the cell is
drafted with a single dye