Fluorene-based fluorescent probes with high two-photon action cross-sections for biological multiphoton imaging applications

Two-photon fluorescence microscopy is a powerful tool for the study of dynamic cellular processes and live-cell imaging. Many commercially available fluorescent probes have been used in multiphoton-based imaging studies despite exhibiting relatively low two-photon absorption cross-section values in the tunability range of ultrafast Ti:sapphire lasers commonly used in multiphoton microscopy imaging. Furthermore, available fluorophores may be plagued with low fluorescence quantum yield and/or photoinstability (i.e., photobleaching) on exposure to the high peak power and photon density provided by the ultrafast laser source. To address the demand for better performing dyes, we prepare fluorophores tailored for multiphoton imaging. These fluorophores are based on the fluorene ring system, known to exhibit high fluorescence quantum yield ( \u3e 0.7) and high photostability. Furthermore, an amine-reactive fluorescent probe for the covalent attachment onto amine-containing biomolecules is also prepared. Epi-fluorescence and two-photon fluorescence microscopy images of H9c2 rat cardiomyoblasts stained with an efficient two-photon absorbing fluorene fluorophore is demonstrated. Additionally, single-photon spectral characteristics of the amine-reactive fluorophore, as well as the two-photon absorption cross sections of its model adduct in solution, and spectral characterization of a bovine serum albumin (BSA) as a model bioconjugate are presented

Prognostic relevance of Centromere protein H expression in esophageal carcinoma

<p>Abstract</p> <p>Background</p> <p>Many kinetochore proteins have been shown to be associated with human cancers. The aim of the present study was to clarify the expression of Centromere protein H (CENP-H), one of the fundamental components of the human active kinetochore, in esophageal carcinoma and its correlation with clinicopathological features.</p> <p>Methods</p> <p>We examined the expression of CENP-H in immortalized esophageal epithelial cells as well as in esophageal carcinoma cells, and in 12 cases of esophageal carcinoma tissues and the paired normal esophageal tissues by RT-PCR and Western blot analysis. In addition, we analyzed CENP-H protein expression in 177 clinicopathologically characterized esophageal carcinoma cases by immunohistochemistry. Statistical analyses were applied to test for prognostic and diagnostic associations.</p> <p>Results</p> <p>The level of CENP-H mRNA and protein were higher in the immortalized cells, cancer cell lines and most cancer tissues than in normal control tissues. Immunohistochemistry showed that CENP-H was expressed in 127 of 171 ESCC cases (74.3%) and in 3 of 6 esophageal adenocarcinoma cases (50%). Statistical analysis of ESCC cases showed that there was a significant difference of CENP-H expression in patients categorized according to gender (<it>P </it>= 0.013), stage (<it>P </it>= 0.023) and T classification (<it>P </it>= 0.019). Patients with lower CENP-H expression had longer overall survival time than those with higher CENP-H expression. Multivariate analysis suggested that CENP-H expression was an independent prognostic marker for esophageal carcinoma patients. A prognostic value of CENP-H was also found in the subgroup of T3~T4 and N0 tumor classification.</p> <p>Conclusion</p> <p>Our results suggest that CENP-H protein is a valuable marker of esophageal carcinoma progression. CENP-H might be used as a valuable prognostic marker for esophageal carcinoma patients.</p

Two-Photon Absorption Spectra Of Fluorene Derivatives

Two-photon absorption (2PA) spectra of fluorene derivatives was presented. A group of fluorene derivatives were synthesized in order to investigate the structure-property relationships of these dyes. Results showed that the overall increase in 2PA illustrated the potential benefit of extending the cyclic conjugation length in fluorene push-pull compounds

Cellular Imaging With High-Performance Two-Photon Absorbing Fluorescent Contrast Agents

A new series of high-performing two-photon absorbing fluorescent contrast agents for cellular imaging were prepared. The spectral properties of these photostable, highly fluorescent, fluorophores, along with twophoton fluorescence imaging of live cells are presented. © 2006 OSA

Resonant enhancement of two-photon absorption in substituted fluorene molecules

© 2004 American Institute of Physics. The electronic version of this article is the complete one and can be found at: http://dx.doi.org/10.1063/1.1770726DOI: 10.1063/1.1770726The degenerate and nondegenerate two-photon absorption (2PA) spectra for a symmetric and an asymmetric fluorene derivative were experimentally measured in order to determine the effect of intermediate state resonance enhancement (ISRE) on the 2PA cross section δ. The ability to tune the individual photon energies in the nondegenerate 2PA (ND-2PA) process afforded a quantitative study of the ISRE without modifying the chemical structure of the investigated chromophores. Both molecules exhibited resonant enhancement of the nonlinearity with the asymmetric compound showing as much as a twentyfold increase in δ. Furthermore, the possibility of achieving over a one order of magnitude enhancement of the nonlinearity reveals the potential benefits of utilizing ND-2PA for certain applications. To model ISRE, we have used correlated quantum-chemical methods together with the perturbative sum-over-states (SOS) expression. We find strong qualitative and quantitative correlation between the experimental and theoretical results. Finally, using a simplified three-level model for the SOS expression, we provide intuitive insight into the process of ISRE for ND-2PA

Predictions of Two Photon Absorption Profiles Using Time-Dependent Density Functional Theory Combined with SOS and CEO Formalisms

Two-photon absorption (2PA) and subsequent processes may be localized in space with a tightly focused laser beam. This property is used in a wide range of applications, including three dimensional data storage. We report theoretical studies of 5 conjugated chromophores experimentally shown to have large 2PA cross-sections. We use the Time Dependent Density Functional Theory (TD-DFT) to describe the electronic structure. The third order coupled electronic oscillator formalism is applied to calculate frequency-dependent second order hyperpolarizability. Alternatively, the sum over states formalism using state-to-state transition dipoles provided by the a posteriori Tamm-Dancoff approximation is employed. It provides new venues for qualitative interpretation and rational design of 2PA chromophores. © 2009 Springer Berlin Heidelberg