Basics of Fluorescence Spectroscopy in Biosciences

Hof, Martin; Fidler, Vlastimil; Hutterer, Rudi

doi:10.1007/3-540-27004-3_1

Cited by 35 publications

(41 citation statements)

References 139 publications

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“…However, there are several drawbacks inherent to this technique, such as photodamage, photobleaching and shallow penetration 35 depths, which can be overcome by utilizing multiphoton processes. 2 Nonlinear optical (NLO) microscopy has emerged as a promising alternative to one-photon fluorescence microscopy for non-invasive 3D imaging of biological tissues. 3 The most common forms of NLO microscopy rely on two nonlinear 40 phenomena: Two-photon excited fluorescence (TPEF) and second-harmonic generation (SHG).…”

Section: Introductionmentioning

confidence: 99%

Thiophene-based dyes for probing membranes

López‐Duarte

Chairatana

et al. 2015

Org. Biomol. Chem.

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We report the synthesis of four new cationic dipolar push-pull dyes, together with an evaluation of their photophysical and photobiological characteristics pertinent for imaging membranes by fluorescence and second harmonic generation. All four dyes consist of an N,N-diethylaniline electron-donor conjugated to a pyridinium electron-acceptor via a thiophene bridge, with either vinylene (-CH=CH-) or ethynylene 10 (-CC-) linking groups, and with either singly-charged or doubly-charged pyridinium terminals. The absorption and fluorescence behavior of these dyes was compared to a commercially available fluorescent membrane stain, the styryl dye FM4-64. The hyperpolarizabilities of all dyes were compared using hyper-Rayleigh scattering at 800 nm. Cellular uptake, localization, toxicity and phototoxicity were evaluated using tissue cell cultures (HeLa, SK-OV-3 and MDA-231). Replacing the central alkene bridge 15 of FM4-64 with a thiophene does not substantially change the absorption, fluorescence or hyperpolarizability, whereas changing the vinylene-links to ethynylenes shifts the absorption and fluorescence to shorter wavelengths, and reduces the hyperpolarizability by about a factor of two. SHG and fluorescence imaging experiments in live cells showed that the doubly-charged thiophene dyes localize in plasma membranes, and exhibit lower internalization rates compared to FM4-64, resulting in 20 less signal from the cell cytosol. At a typical imaging concentration of 1 µM, the doubly-charged dyes showed no significant light or dark toxicity, whereas the singly-charged dyes are phototoxic even at 0.5 µM, and toxic in the dark at concentrations above 5 µM. The doubly-charged dyes showed phototoxicity at concentrations greater than 10 µM, although they do not generate singlet oxygen, indicating that the phototoxicity is type I rather than type II. Our data demonstrate that the doubly-charged thiophene dyes 25 are more effective than FM4-64 as nonlinear optical imaging agents for live cells.

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Section: Introductionmentioning

confidence: 99%

Thiophene-based dyes for probing membranes

López‐Duarte

Chairatana

et al. 2015

Org. Biomol. Chem.

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“…Although there has been rapid growth in the number of routinized fluorescence applications, the principles have remained the same. 2) Fluorescence spectrometry is a conventional but highly sensitive analytical method. The synthesis and the biological applications of fluorescent compounds have previously been investigated.…”

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confidence: 99%

Electro-Organic Synthesis and Characterization of New Dihydroxybenzene Dinitrile Derivatives with Fluorescent Properties

Moghaddam

Dinarvand

Latifi

et al. 2008

CHEMICAL & PHARMACEUTICAL BULLETIN

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“…Fluorescence spectroscopy studies the fluorescence light emission as function of the wavelength. To date, fluorescence spectroscopy is already integrated in an extensive range of applications in, amongst others, the biochemical, medical and petrochemical industry [1][2][3][4][5]. In biology and chemistry, it facilitates the study of proteins and the tracking of biochemical reactions.…”

Section: Introductionmentioning

confidence: 99%

Improving Food Safety by Using One- and Two-Photon- Induced Fluorescence Spectroscopy for the Detection of Mycotoxins

Smeesters¹,

Meulebroeck²,

Thienpont³

2016

Applications of Molecular Spectroscopy to Current Research in the Chemical and Biological Sciences

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The presence of mycotoxins in food products is a major worldwide problem. Nowadays, mycotoxins can only be detected by the use of sample-based chemical analyses. Therefore, we demonstrate the use of one-and two-photon-induced fluorescence spectroscopy for the non-destructive detection of mycotoxins in unprocessed food products. We first explain our optical set-up, which is able to measure the localized oneand two-photon-induced fluorescence spectra. Following, as a case study, the detection of aflatoxin in maize kernels is discussed. We present our research methodology, from the characterization of the fluorescence of pure aflatoxin, to the study of the one-and two-photon-induced fluorescence spectra of maize kernels and the development of an optical detection criterion. During both one-and two-photon-induced fluorescence processes, the fluorescence of the aflatoxin influences the intrinsic fluorescence of the maize. Based on the fluorescence spectrum between 400 and 550 nm, a detection criterion to sense the contaminated kernels is defined. Furthermore, we successfully monitored the localized contamination level on the kernel's surface, showing both contaminated kernels with a high contamination in a limited surface area (a few square millimetres) and kernels with a low contamination spread over a large surface area (up to 20 mm 2 ). Finally, the extensibility of our research methodology to other fluorescent mycotoxins is discussed.

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Basics of Fluorescence Spectroscopy in Biosciences

Cited by 35 publications

References 139 publications

Thiophene-based dyes for probing membranes

Thiophene-based dyes for probing membranes

Electro-Organic Synthesis and Characterization of New Dihydroxybenzene Dinitrile Derivatives with Fluorescent Properties

Improving Food Safety by Using One- and Two-Photon- Induced Fluorescence Spectroscopy for the Detection of Mycotoxins

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