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Appl Spectrosc. 2016 Jul;70(7):1195-201. doi: 10.1177/0003702816652363. Epub 2016 May 26.

Using Pyridinium Styryl Dyes as the Standards of Time-Resolved Instrument Response.

Author information

1
State Key Laboratory of Precision Spectroscopy, East China Normal University, Shanghai, China.
2
State Key Laboratory of Precision Spectroscopy, East China Normal University, Shanghai, China sjzhang@phy.ecnu.edu.cn.
3
JILA, National Institute of Standards and Technology (NIST), University of Colorado, Boulder, Colorado, USA.

Abstract

In this paper, two pyridinium styryl dyes, [2-(4-dimethylamino-phenyl)-vinyl]-1-methylpyridinium iodide (DASPMI), were synthesized and characterized by steady state fluorescence spectroscopy as well as picosecond and femtosecond time-resolved fluorescence spectroscopies. Both dyes exhibit large Stokes shifts and fluorescence decays equivalent to the instrument response function (IRF) standards employed in time-correlated single-photon counting. Due to their styryl and pyridinium moieties, DASPMIs have higher peak fluorescence intensity and shorter excited-state lifetimes than iodide ion-quenched fluorophores. The fluorescence lifetimes of o-DASPMI and p-DASPMI were measured to be 6.6 ps and 12.4 ps, respectively. The fluorescence transients of these DASPMIs were used as the IRFs for iterative reconvolution fitting of the time-resolved fluorescence decay profiles of Rhodamine B (RhB), sulforhodamine B (SRB), and the SRB-SRB2m RNA aptamer complex. The quality of the fits employing the DASPMI-derived IRFs are consistently equivalent to those employing IRFs obtained from light scattering. These results indicate that DASPMI-derived IRFs may be suited for a broad range of applications in time-resolved spectroscopy and fluorescence lifetime imaging microscopy (FLIM), especially in the visible emission range.

KEYWORDS:

Fluorescence quenching; instrument response function (IRF); pyridinium styryl dyes; time-resolved fluorescence

PMID:
27231333
DOI:
10.1177/0003702816652363

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