[Intensity loss of two-photon excitation fluorescence microscopy images of mouse oocyte chromosomes].

As an optical microscope with high resolution, two-photon excitation (TPE) fluorescence microscope is widely used in noninvasive 3D optical imaging of biological samples. Compared with confocal laser scanning microscope, TPE fluorescence microscope provides a deeper detecting depth. In spite of that, the image quality of sample always declines as the detecting depth increases when a noninvasive 3D optical imaging of thicker samples is performed. Mouse oocytes with a large diameter, which play an important role in clinical and biological fields, have obvious absorption and scattering effects. In the present paper, we performed compensation for two-photon fluorescence images of mouse oocyte chromosomes. Using volume as a parameter, the attenuation degree of these chromosomes was also studied. The result of our data suggested that there exists a severe axial intensity loss in two-photon microscopic images of mouse oocytes due to the absorption and scattering effects. It is necessary to make compensation for these images of mouse oocyte chromosomes obtained from two-photon microscopic system. It will be specially needed in studying the quantitative three-dimensional information of mouse oocytes.

[Improved data processing method of fluorescence correlation spectroscopy].

By the unique ability of measuring concentration and diffusion coefficient, fluorescence correlation spectroscopy (FCS) has kept enlarging and deepening its application fields which need measuring physicochemical properties in complex mixtures. However, the classical data processing method in FCS generally induces errors in the fitting parameters, and the intuitionistic information in the chart is little or confused. Aiming at the above problems, we tried a new data processing method with differential treatment to find out whether it has multi-fluorescent-components and estimate the parameters intuitionally in the chart. The differential curves of treated autocorrelation curves have various positions, depths and full widths at half depth of the trough, by which the characteristic diffusion time and the number of the fluorescent components are decided. This improved processing method provides help for the FCS measurement in complex environment of life.

[Application of two-photon excitation fluorescence imaging to real-time investigation of mouse preimplantation embryo].

The blue shift of a two-photon excitation absorption peak allows the application of a single wavelength to the simultaneous excitation of several fluorochromes with disparate emission characteristics. An output at 730 nm of a mode-locked femtosecond Ti-sapphire laser was used to excite four different commonly used fluorochromes, namely Hoechst 33342, Fluo-4, PI and Indo-1, and characteristic fluorescence images were obtained using (455 +/- 15) nm, (540 +/- 15)nm, (580 +/- 16) nm and (500 +/- 15) nm filters respectively. An approach to exciting with a single wavelength, staining with two different fluorochromes, and collecting fluorescence in two separate channels was employed to study mouse preimplantation embryos by 3D and 4D real-time imaging. Combined with the merits of two-photon excitation fluorescence imaging, such as better penetration, less photon-damage, and higher signal-to-noise ratio, this approach was supposed to be a novel multi-parameter investigating tool for mouse preimplantation embryos development study.

[Measurement of methane 13C/12C isotope ratio of slurry gas from oil wells by laser frequency-modulation absorption spectroscopy].

In this paper, we determined the ratio of C13/C12 of methane in slurry gas from oil wells by the method of laser frequency modulation absorption spectroscopy that has many advantages such as high precision, high spectroscopic resolution and rapidly and simply dealing with samples. This technique provided us with a subsidiary method for oil-gas source exploration. The method of laser frequency modulation absorption spectroscopy is based on the method of laser frequency absorption spectroscopy and is more accurate. And this method can be expanded to apply to others aspects easily. We only need change other appropriate laser and select right absorption peak, then we can determined the ration and concentration of diverse gases.

[Fluorescence fluctuation spectroscopy with strong background fluorescence--a Monte Carlo approach].

Fluorescence fluctuation spectroscopy is a method in which fluorescence fluctuations arising from a very small sample volume are analysed to obtain brightness, diffusion coefficient and concentration of particles. Using Monte Carlo simulation, the influences of background autofluorescence and noises on fluorescence fluctuation spectroscopy are studied. Results show that a two-component photon counting histogram can effectively remove the effects due to background autofluorescence from low brightness and high concentration components and uniform noise. The result will help to bring applications of fluorescence fluctuation spectroscopy to intracellular environment.

Capillary electrophoresis and circular dichroism study of trichosanthin and its mutants.

The type-I ribosome-inactivating protein trichosanthin (TCS) has a broad spectrum of biological and pharmacological activities, including abortifacient, anti-tumor and anti-human-immunodeficiency-virus (anti-HIV). In this study, circular dichroism (CD) and capillary electrophoresis were used for the first time to study TCS and its two TCS mutants of Y55G TCS (tyrosine 55 converted to glycine) and FYY140-142GSA TCS (tripeptide phenylalanine-tyrosine-tyrosine 140-142 converted to glycine-serine-alanine). The results indicated that the substitution of amino acids changed the secondary structures and the hydrophobility of TCS. Moreover, both Y55G TCS and FYY140-142GSA TCS demonstrated attenuated cytotoxicity and reactive oxygen species (ROS) production in human choriocarcinoma cells (JAR cells) as compared to natural TCS and wild-type TCS. Our results demonstrated the cytotoxicity of TCS on JAR cells and TCS-induced production of ROS might be TCS-conformational related, suggesting that CD and capillary electrophoresis study might throw new insight into the anti-tumor and anti-HIV mechanism of TCS.