Dye-induced fluorescence of tumour cells: photochemical action spectra (original) (raw)
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Photodynamic activity of different dyes
Laser Physics, 2007
Photodynamic therapy (PDT) is a technique for inducing tissue damage with light irradiation of a drug selectively retained in malignant tissue. Many kinds of compounds are known with photosensitizing properties including dyes, drugs, cosmetics, chemicals, and many natural substances. There are different classes of sensitizers used for medical purposes such as tetrapyrroles (porphyrins and derivatives, chlorophyll, phylloerythrin, phthalocyanines), tricyclic dyes with different meso-atoms (acridine orange, proflavine, riboflavine, methylene blue, fluorescein, eosine, erythrosine, rose bengal), and furocoumarins (psoralen and its methoxyderivatives xanthotoxin, bergaptene). In this work, we performed one comparative cytotoxic study of the photodynamic activity presented by tricyclic dyes (methylene blue, fluorescein and erythrosine) and the commercial Russian photosensitizer Photogem® (hematoporphyrin derivative). For this purpose, three cell lines were used: HEp-2 (tumor cells), VERO and McCoy (nontumor cells), and a yeast strain. The wavelength used for irradiation was 630 nm, the same as used in PDT for medical purposes, since it is in the therapeutic window, i.e., where light can penetrate more into the tissues. The results suggest that Photogem® is more cytotoxic and more photocytotoxic than the studied tricyclic dyes in nontumor and tumor cells. These dyes present less cytotoxicity (around half) in normal cells (dark and light) than in tumor cells. In the experiments with microorganisms, methylene blue presented a better photodynamic effect than Photogem®. These results can be explained by the fact that it is more difficult for Photogem® to penetrate in microorganism membranes than mammalian cell membranes. As for Photogem®, these tricycle dyes present a higher cytotoxicity in tumor cells. These data suggest that methylene blue can be an option in photodynamic inactivation as well as in photodynamic therapy, mainly for superficial lesions.
Uptake and accumulation of the vital dye hydroethidine in neoplastic cells
Journal of Histochemistry and Cytochemistry, 1986
Hydroethidine, a reduced form of ethidium bromide, was used as a vital dye in fluorescence assays that allowed visual and semiquantitative monitoring of dye uptake and accumulation by fluorescence microscopy, flow cytometry, image analysis, and microfluorimetry. The excitation and emission filters were chosen to detect hydroethidine and exdude ethidium. Microscopically, there were differences in fluorescence intensities and fluorescence patterns among various tumor cell lines. The fluorescence pattern varied from homogeneous blue in the cytoplasm to blue plus brilliant packets of bluish-white distributed in the cytoplasm. Nudear staining varied from brown to reddish orange fluorescence. These differences were confirmed by flow cytometry and image analysis. A preliminary survey of van
Bioconjugate Chemistry, 2006
In vivo optical imaging to enhance the detection of cancer during endoscopy or surgery requires a targeted fluorescent probe with high emission efficiency and high signal-to-background ratio. One strategy to accurately detect cancers is to have the fluorophore internalize within the cancer cells permitting nonbound fluorophores to be washed away or absorbed. The choice of fluorophores for this task must be carefully considered. For depth of penetration, near-infrared probes are ordinarily preferred but suffer from relatively low quantum efficiency. Although green fluorescent protein has been widely used to image tumors on internal organs in mice, green fluorescent probes are better suited for imaging the superficial tissues because of the short penetration distance of green light in tissue and the highly efficient production of signal. While the fluorescence properties of green fluorophores are well-known in vitro, less attention has been paid to their fluorescence once they are internalized within cells. In this study, the emission efficiency after cellular internalization of four common green fluorophores conjugated to avidin (Av-fluorescein, Av-Oregon green, Av-BODIPY-FL, and Av-rhodamine green) were compared after each conjugate was incubated with SHIN3 ovarian cancer cells. Using the lectin binding receptor system, the avidin-fluorophore conjugates were endocytosed, and their fluorescence was evaluated with fluorescence microscopy and flow cytometry. While fluorescein demonstrated the highest signal outside the cell, among the four fluorophores, internalized Av-rhodamine green emitted the most light from SHIN3 ovarian cancer cells both in vitro and in vivo. The internalized Av-rhodamine green complex appeared to localize to the endoplasmic vesicles. Thus, among the four common green fluorescent dyes, rhodamine green is the brightest green fluorescence probe after cellular internalization. This information could have implications for the design of tumor-targeted fluorescent probes that rely on cellular internalization for cancer detection.
Synthesis of dye conjugates to visualize the cancer cells using fluorescence microscopy
Applied Optics, 2014
The clinical diagnosis of most cancers is based on evaluation of histology microscopic slides to view the size and shape of cellular nuclei and morphological structure of tissue. To achieve this goal for in vivo and in-deep tissues, near infrared dyes-bovine serum albumin and immunoglobulin G conjugates were synthesized. The spectral study shows that the absorption and fluorescence of the dye conjugates are in the "tissue optical window" spectral ranges between 650 and 900 nm. The internalization and pinocytosis of the synthesized compounds were investigated at cell level using fluorescence microscopy to obtain the optimal concentration and staining time.
Journal of Photochemistry and Photobiology B: Biology, 1995
Steady state and time-resolved fluorescence spectroscopy were employed to study the fluorescence from non-metastatic, metastatic and non-tumorigenic cell lines from different species. Excitations at 310 nm and 350 nm were used to monitor tryptophan and reduced nicotinamide adenine dinucleotide (NADH) fluorescence respectively. Subtle and consistent differences were observed between different categories of cell lines. It was found that the tryptophan to NADH fluorescence intensity ratio is higher in metastatic cell lines than in non-metastatic and normal cell lines. The fluorescence decay of the tryptophan residue in different cell lines was best described by triple exponential kinetics, whereas lhe NADH fluorescence decay was best described by mainly double and, in some cases, triple exponential kinetics. The average fluorescence lifetimes for tryptophan were in the range 2.5-3.7 ns. The average lifetime of NADH was lower (by a factor of approximately three) in metastatic cells than in non-metastatic ceils and this finding is consistent for cell lines from different origins (rat or human). Correcting the Iluorescence intensity for the average fluorescence lifetime of each species and for the volume of each cell line, it was shown that the ~'oncentrations of tryptophan and NADH are consistently higher in malignant metastatic cancer cells than in non-metastatic cells.
Cell Staining by Novel Derivatives of Fluorescent Rhodamine Dyes
Two novel precursors of fluorescent dyes (PFD813 and PFD814) have been studied for their ability to photo-activation, transfer across the biomembrane and cells staining. The fluorescent dyes Rho813 and Rho814 formed by photo-activation of their precursors PFD813 and PFD814 inside cells were used for the optical detection of particular features in vitro (HaCat cells, human epithelial carcinoma A431, epidermoid carcinoma of the cervix HeLa and chinese hamster ovary CHO cells). One of the possibilities to visualize and track the pathways of macromolecules or organelles in a " living " cell is to monitor them after staining with these PFDs during the real time measurements. A bright fluorescent signal from the photoactivated dye molecules inside the small spot in the cell can be monitored during their movement into the cell dark region (where the dye was not activated and did not fluoresce). The obtained data are important for further application of these precursors of the fluorescent dyes (" caged " dyes) for microscopic probing of biological objects.
Reactive dyes for living cells: Applications, artefacts, and some comparisons with textile dyeing
Coloration Technology, 2021
An inclusive chemical definition of “reactive” dyeing of textiles is introduced, encompassing the CI Azoic, CI Mordant, CI Reactive, CI Sulphur and CI Vat dye application classes. Such reactive dyeing increases fibre retention of dye and makes application practically possible. The analogous application of dyes and fluorescent probes as microscopic stains in biology and medicine is outlined, focussing on using reactive fluorescent probes with living cells. Parallels with textile dyeing are noted, eg, enhanced probe retention and facilitation of probe application. However, the primary purpose of using reactive probes with live cells is detection of properties of biological systems: to identify biological structures and chemical/biochemical contents; assess biological functions and physicochemical properties; and determine changes in locations of cells and cell components. Problems occurring with such probes are outlined, particularly the problematic character of many standard protocol...
Specific fluorescent tracers. Imaging and applications for photodynamic therapy
Comptes Rendus Biologies, 2002
Our main objective is to enlarge the fluorescence use in biosciences, with especially the photodynamic therapy (PDT) used for cancer treatment as one of the target applications. Meta-tetra(hydroxyphenyl)chlorin (m-THPC) is a second-generation photosensitiser, applied in photodynamic therapy. The localisation of this sensitiser as well as its induced cell death mechanisms in human breast cancer cells (MCF-7 and its resistant subline MCF-7 DXR , DXR: doxorubicin) were evaluated using fluorescence microscopy. In addition, we will present two additional routes, whose aims are to create new features to respond to the PDT questioning: firstly, the synthesis of fluorescent tracers, with a particular attention to the presence of hydrophilic groups (glucosamine ring) on the basic fluorophore structure to orientate the localisation of the probe and, secondly, the use of scanning near-field optical microscopy to reach a better resolution for the fluorescence microscopy analysis. To cite this article: M.-H. Teiten et al., C. R. Biologies 325 (2002) 487-493. © 2002 Académie des sciences / Éditions scientifiques et médicales Elsevier SAS fluorescence microscopy / fluorescent tracers / photodynamic therapy / biological cells / scanning near-field optical microscopy Résumé-Traceurs fluorescents spécifiques. Imagerie et applications en thérapie photodynamique. Notre objectif est d'accroître l'utilisation de la fluorescence pour les biosciences, avec comme application la thérapie photodynamique (PDT) utilisée pour le traitement de certains cancers. La méta-tétra(hydroxyphényl)chlorine (m-THPC) est un photosensibilisant de seconde génération utilisé en PDT. Sa localisation dans des cellules humaines de cancer du sein (MCF-7 et sa lignée résistante MCF-7 DXR) a été étudiée par microscopie de fluorescence, afin d'appréhender les cibles impliquées dans la photo-inactivation des cellules et d'évaluer les mécanismes de mort cellulaire induits. De plus, deux nouvelles voies de recherche, qui ont pour but d'améliorer les études en PDT, seront brièvement décrites. Il s'agit, d'une part, de la synthèse de traceurs fluorescents modifiés par la présence d'un cycle hydrophile glucosamine, de manière à orienter leur localisation à l'échelle cellulaire, et, d'autre part, de l'utilisation de la microscopie optique en champ proche pour atteindre, avec une meilleure résolution, les sites de fixation des traceurs.