Valeriy M. Yashchuk

A Monomethine Cyanine Dye Cyan 40 for Two-photon–excited Fluorescence Detection of Nucleic Acids and Their Visualization in Live Cells¶

Abstract Monomethine cyanine dye 4-((1-methylbenzothiazolyliliden-2)methyl)-1,2,6-trimethylpyridinium perchlorate (Cyan 40) was investigated as a two-photon–excited fluorescence probe for nucleic acids (NA). Cyan 40 has been shown to demonstrate efficient two-photon–excited fluorescence in the presence of NA in vitro in contrast to solutions without NA. Two-photon confocal laser scanning microscopy (TPCLSM) and two-photon laser scanning microspectrofluorometry were used to check the possibility of using Cyan 40 as two-photon–excited fluorescence label for NA in living cells. Study of dye effect on viability of cells was also carried out. We ascertained that Cyan 40 is a cell-permeant dye, manifesting efficient two-photon–excited fluorescence when bound to NA in living cells, without any significant influence on viability of cells. TPCLSM images obtained from stained cells indicate preferential RNA staining by Cyan 40 compared with DNA.

Interaction of cyanine dyes with nucleic acids. XVII. Towards an aggregation of cyanine dyes in solutions as a factor facilitating nucleic acid detection

Spectral properties of newly synthesized cyanine dyes, namely 1-[6-(4-[6-[2,6-dimethyl-4-(3-methyl-2,3-dihydro-1,3-benzothiazol- 2-ylidenmethyl)-1-pyridiniumyl]hexanoyl]piperazino)-6- oxohexyl]-2,6-dimethyl-4-(3-ethyl-2,3-dihydro-1,3-benzothiazol+ ++-2-ylidenmethyl)pyridinium (K-6) (bichromophoric dye) and 1-[5-di(3-[5-[2,6-dimethyl-4-(3-methyl-2,3-dihydro-1,3-benzothiazol++ +-2-ylidenmethyl)-1-pyridiniumyl]pentylcarboxamido]pro pyl) carbamoylpentyl]-2,6-dimethyl-4-(3-methyl-2,3-dihydro-1,3-benzo thiazol-2-ylidenmethyl) pyridinium (K-T) (trichromophoric dye) in solutions in the presence of and without deoxyribonucleic acid (DNA) were studied within a wide concentration range. It has been established that absorption, as well as fluorescence of investigated dye solutions, without DNA are mainly determined by H-aggregates of dye molecules. On the contrary, the fluorescence of dye solutions in the presence of DNA gives an intrinsic dye molecular fluorescence. H-aggregates are broken because of binding dye molecules with DNA. It has been suggested that both K-T and K-6 molecules bind mainly with DNA via the interaction of two chromophores. As the ratio of the number of dye molecules to that of DNA base pairs increases with an increase in dye concentration, a formation of dye molecule H-aggregates on DNA molecules are observed. Such aggregates have a different structure than those formed in the solutions without DNA. On the grounds of the data obtained, it is concluded that it is possible to use a dye aggregation capable of obtaining higher values for fluorescence enhancement of the DNA stains.

Self-organization C60 nanoparticles in toluene solution

The structure of fullerene C60 in toluene solution was investigated both experimentally and theoretically in detail. C60 was found to aggregate slowly even in fairly dilute solution concentrations ranging from 0.18 to 0.78 g/l at room temperature. The electronic absorption spectra obtained testify to the mainly molecular character of absorption. The aggregation kinetics results indicated that the structure of C60 aggregates could be described as a fractal system. The numerical calculations predicted the formation of stable fullerene (C60)N clusters in toluene solution with sizes >1.2 nm and aggregation number N≥3.

Biophysical studies of fullerene-based composite for bio-nanotechnology

Abstract The UV absorption and fluorescence spectra of biological samples and the DNA structural state in cells under X-ray irradiation in the presence of the fullerene-containing composite, synthesized on the basis of aminopropylaerosil, were investigated and analysed in detail. The significant pro-oxidant effect of the fullerene-containing composite at the low concentration of fullerene C60 (10−5 M) was revealed.

Fluorescence of Styryl Dyes-DNA Complexes Induced by Single- and Two-Photon Excitation

The series of novel monomer and homodimer styryl dyes based on (p-dimethylaminostyryl) benzothiazolium residues were synthesized and studied as possible fluorescent probes for nucleic acids detection. Spectral-luminescent and spectral-photometric properties of obtained dyes in the unbound state and in DNA presence were studied. Fluorescence emission induced by two-photon excitation of dye-DNA complexes in aqueous buffer solution was registered. Two-photon absorption cross section values of the studied dyes in DNA presence were evaluated.

Styryl dyes as two-photon excited fluorescent probes for DNA detection and two-photon laser scanning fluorescence microscopy of living cells.

Spectral-fluorescent properties of benzothiazole styryl monomer (Bos-3) and homodimer (DBos-21) dyes in presence of DNA were studied. The dyes enhance their fluorescence intensity in 2–3 orders of magnitude upon interaction with DNA. Studied styrylcyanines in DNA presence demonstrate rather high values of two-photon absorption (TPA) cross-section, which are comparable with the values of TPA cross section of the rhodamine dyes. An applicability of the styrylcyanines as probes for the fluorescence microscopy of living cells was studied. It was shown that both dyes are cell-permeable but homodimer dye DBos-21 produces noticeably brighter staining of HeLa cells comparing with monomer dye Bos-3. Molecules of DBos-21 initially bind to the nucleic acids- containing cell organelles (presumable mitochondria) and are able to penetrate into the cell nucleus. Thus, homodimer styryl DBos-21 dye is viewed as efficient stain for single-photon and two-photon excitation fluorescence imaging of living cells.

Electronic Excitation Energy Transfer in DNA. Nature of Triplet Excitations Capturing Centers

Spectral properties, energy structure, and electronic processes in DNA, poly(dAdT)2, d(CCCGGGTTTAAA), d(ATC), d(AT), dGMP, dAMP, dCMP, dTMP were studied. The DNA lowest excited electronic singlet and triplet levels are connected with guanine and adenine bases, correspondingly. The DNA fluorescence is close to a linear combination of the guanine and cytidine emissions and connected, to a greater extent, with the singlet energy transfer to these bases. The origin of the DNA phosphorescence is associated with AT-sequences that are traps of triplet excitations in DNA. The triplet energy transfer to these centers and adenine bases (that are most photostable) leads to the DNA self-protection against a damage induced by electronic excitations of DNA.

Nonradiative deactivation of the electronic excitation energy in cyanine dyes: influence of binding to DNA.

The processes of nonradiative deactivation of electronic excitation energy in cyanine dyes determine their quantum yield. Because of that, the study of the influence of cyanines binding to DNA on these processes can provide information on the causes leading to the cyanines fluorescence intensity enhancement in the presence of DNA. In the presented paper, the activation energies of nonradiative degradation of electronic excitation, quantum yields and rate constants of nonradiative transitions of several cyanines in free state and in the presence of DNA were established and compared. The mechanisms of nonradiative deactivation of dye excitation energy were discussed.

Migrating electronic excitations in π-electron-containing polymers

Abstract The results of the investigations of spectral properties and excitation energy transfer in π-electron-containing polymers carried out in Kyiv University are briefly reviewed. The peculiarities of fluorescence, delayed fluorescence, singlet and triplet exciton propagation, mechanisms of elementary acts of singlet and triplet energy transfer are examined and analyzed. Some proofs of vibrational excitons in π-electron-containing polymers are presented.

Multifunctional Macromolecules and Structures as One-Way Exciton Conductors

Abstract The first steps in the design, investigations and possible applications of multifunctional macromolecules exhibiting one-way direct exciton conductivity are presented. Some problems that appeared during the formation of these systems are discussed.