口腔常见致龋菌荧光光谱特征的初步研究

目的采用三维荧光光谱分析技术对口腔常见致龋菌荧光光谱特征进行初步分析。方法选择口腔常见致龋菌变形链球菌及远缘链球菌,对其进行复苏和培养,运用三维荧光光谱分析技术对其菌液进行荧光学光谱检测。结果变形链球菌及远缘链球菌的三维荧光光谱图相似,均出现2个荧光峰,最佳激发波长分别位于230nm和280nm,最佳发射波长相同,均为340nm。结论成功获得口腔常见致龋菌(变形链球菌及远缘链球菌)的固有荧光三维光谱图,为致龋菌荧光评价技术的应用提供参考。     

Dynamics of the fluorescence properties of pyrene residues appended to oligonucleotide hybridization probes.

The dynamic and static properties of the fluorescence of a pyrene-introduced oligonucleotide 16 mer and its hybrid with a target 32 mer. Their fluorescence quantum yields (< 1%) were much weaker than that of unsubstituted pyrene and their fluorescence lifetime of the major decay components were less than 1 ns. The rapid fluorescence quenching was due to the interaction between the fluorophore and bases in the oligonucleotides. The fluorescence of pyrene was quenched efficiently by TMP and slightly by AMP. The quenching by CMP and GMP were the intermediate case.     

Specificity of antinuclear autoantibodies in diseases with systemic destruction of connective tissue]

By using the fluorescent antibody technique several patterns of antinuclear antibodies (ANA) were detected in the systemic connective tissue diseases, as well as in some other diseases and in healthy donors, depending on the character of nuclear fluorescence. Homogeneous nuclear fluorescence, homogeneous fluorescence free of nucleaolar fluorescence, ring-shaped fluorescence, lumpy fluorescence, selective nuclealar fluorescence, nuclear fluorescence in the form of long fine plexiform bands associated with fluorescence in the nuclear membrane region. The latter ANA pattern was found only in several forms of lupus erythematosus, and this fluorescence was different from the previously reported "reticular" and "filamentous" patterns.     

A fluorescence study of egg white riboflavin-binding protein

1. Denaturation of riboflavin-binding protein (RBP) by guanidine hydrochloride (Gu-HCl) was investigated by measruing the fluorescence of the protein. The denaturation-renaturation processes of RBP by Gu-HCl were fully reversible. The apo-RBP fluorescence had an emission maximum at 343 nm in the absence of Gu-HCl, and at 350 nm in the presence of 4M Gu-HCl, which completely denatured the protein. The relative fluorescence yield of apo-RBP in the presence of 4 M Gu-HCl was about 170% of that in the absence of Gu-HCl. The affinity of native apo-RBP for riboflavin was very strong, while riboflavin was not bound to the denatured form. The equilibrium system of apo-RBP and riboflavin in solutions containing Gu-HCl at various concentrations was analyzed by measuring riboflavin fluorescence. 2. The quenching of apo-RBP fluorescence, probably the fluorescence of tryptophanyl residues, by iodide anions and cesium cations was measured. The fluorescence of apo-RBP in the presence of 4 M Gu-HCl was quenched considerably by iodide and cesium, and Stern-Volmer plots were linear. However, the fluorescence of native apo-RBP was scarcely quenched by iodide or cesium. This suggested that tryptophanyl residues buried inside apo-RBP were responsible for most of the tryptophanyl fluorescence of native apo-RBP.     

Investigation of the interactions of quercetin and morin with trypsin

The interactions of quercetin and morin with trypsin were investigated by UV–vis absorption, fluorescence, synchronous fluorescence and three‐dimensional fluorescence spectra techniques under physiological pH 7.40. Quercetin and morin effectively quenched the intrinsic fluorescence of trypsin via static quenching. The process of binding quercetin and morin on trypsin was a spontaneous molecular interaction procedure. The binding constants and thermodynamic parameters at two different temperatures, the binding locality and the binding power were obtained. The conformation of trypsin was discussed by synchronous and three‐dimensional fluorescence techniques. Copyright © 2009 John Wiley & Sons, Ltd.     

Effects of chill stress on prompt and delayed chlorophyll fluorescence from leaves

This paper describes the utilization of a portable solid state device for the simultaneous measurement of prompt and delayed fluorescence transients in leaves from a variety of species subjected to temperature lowering. The induction transients of the two phenomena were not identical as the peak in prompt fluorescence yield always preceded that of delayed fluorescence. Temperature lowering delayed the occurrence of peak fluorescence, increased prompt fluorescence yield, decreased delayed fluorescence yield, and caused the occurrence of a new, more rapid delayed fluorescence transient. Leaves from all species had qualitatively the same type of induction curves although the response to temperature differed between species. The delayed fluorescence yield of chill-sensitive species was reduced to a greater extent than that of chill-insensitive species. Cold hardening leaf material did not greatly change the fluorescence response to temperature lowering. Arrhenius plots showed a linear relationship between delayed fluorescence yield and temperature. There were no breaks that would suggest membrane lipid phase changes. The data indicate that thylakoid membranes of chill-sensitive species are less capable of maintaining a light-induced high energy state at low temperatures than are thylakoid membranes of chill-resistant species.     

On-line fluorescence profile of aerobic sludge digestion

An online fluorometer designed for following intracellular NAD(P)H was used to monitor aerobic sludge digestion experiments. The fluorescence showed an initial rise to a high plateau, a sharp decline after staying at the plateau for 20-60 h, and a trailing very slow decrease. The characteristic fluorescence profile was shown to result mainly from the solids-associated fluorescence, after ruling out other factors such as pH, temperature, and supernatant fluorescence. The fluorescence profile was, however, not a mere result of the decreasing solids concentration. The varying sludge viability and population composition (e.g., the decay of heterotrophs and the increasing fraction of nitrifiers) played important roles. The fluorescence profile correlated well with the profile of the viable heterotrophic cell number concentration evaluated with TSB-agar plates. The initial increase of the number concentration was attributed to the growth of multiple small bacteria from the lysate of each large microorganism, which was demonstrated in the experiments with baker's yeast as the starting culture for digestion. The fluorescence profiles observed in the yeast experiments were similar to those in the sludge experiments. Responding to glucose additions and the switch from aerobic to anaerobic conditions, the yeast systems showed typical step increases of fluorescence as expected from the change of NAD(P)H level associated with heterotrophic metabolism. However, no such fluorescence responses were detectable in the sludge digestion systems. NAD(P)H were thus uncertain to be responsible for the online fluorescence observed. Nonetheless, the initial fluorescence plateau corresponded to the period of rapid digestion and, for the plant studied, the EPA regulation criteria of VSS reduction >38% and/or SOUR <1.5 mg of O(2) (g of TS)(-)(1) h(-)(1) were satisfied at the end of the plateau. The online fluorescence provides an effective means of monitoring the aerobic sludge digestion process.     

Accurate determination of DNA content in single cell nuclei stained with Hoechst 33258 fluorochrome at high salt concentration

In an attempt to achieve accurate quantification of DNA levels in cell nuclei, we studied the influence of salt concentration on the fluorescence of cell nuclei complexed with Hoechst-33258 (Hoe) fluorochrome. The fluorescence of cell nuclei was compared with that of extracted DNA as well as that of nucleosome core. Conformational changes in these complexes were examined by measuring both fluorescence anisotropy and fluorescence lifetime in the nanosecond region. The results showed that the fluorescence of DNA-Hoe was quenched by the nucleosomal structure, there being an associated increase in anisotropy and a decrease in the fluorescence lifetime; however, the fluorescence was restored to the original level by the addition of a high concentration of NaCl, CsCl, or LiCl. The reduction in fluorescence may have been due to loss of fluorescence energy caused by collision of the fluorophore with histones in the nucleosome. The addition of 1 M NaCl to the medium used for staining with Hoe greatly stabilized the fluorescence of DNA in cell nuclei. The DNA content of individual cell nuclei was determined by comparing the fluorescence of these nuclei with that of a standard DNA solution. For lymphocytes and liver ploidy cells, reasonably accurate values were obtained by applying the present method.     

Temperature dependence of tryptophan fluorescence lifetime in aqueous glycerol and trehalose solutions

The temperature dependences of tryptophan fluorescence decay kinetics in aqueous glycerol and 1 M trehalose solutions were examined. The fluorescence decay kinetics were recorded in the spectral region of 292.5–417.5 nm with nanosecond time resolution. The kinetics curves were approximated by the sum of three exponential terms, and the spectral distribution (DAS) of these components was determined. An antisymbatic course of fluorescence decay times of two (fast and medium) components in the temperature range from –60 to +10°C was observed. The third (slow) component showed only slight temperature dependence. The antisymbatic behavior of fluorescence lifetimes of the fast and medium components was explained on the assumption that some of the excited tryptophan molecules are transferred from a short-wave-length B-form with short fluorescence lifetime to a long-wavelength R-form with an intermediate fluorescence lifetime. This transfer occurred in the indicated temperature range.     

Evaluation of binding and thermodynamic characteristics of interactions between a citrus flavonoid hesperitin with protein and effects of metal ions on binding

The mechanism of interaction of a non-glycosidic citrus flavonoid, hesperitin (HES) with bovine serum albumin (BSA) was studied by UV-vis absorption, fluorescence, FT-IR, circular dichroism, fluorescence anisotropy and synchronous fluorescence spectroscopy in phosphate buffer of pH 7.4. Fluorescence data revealed that the fluorescence quenching of BSA by HES was the result of the formed complex of HES-BSA. The binding constants and thermodynamic parameters at four different temperatures, the location of binding, and the nature of binding force were determined. The hydrogen bonds interactions were found to be the predominant intermolecular forces to stabilize the complex. The conformation of BSA was discussed by synchronous fluorescence and CD methods. The alterations of protein secondary structure upon complexation with HES were evident from the gradual decrease in α-helicity. The distance between the donor (BSA) and acceptor (flavonoid) was calculated from the fluorescence resonance energy transfer and found to be 1.978 nm. Common ions viz., Zn(2+), K(+), Cu(2+), Ni(2+), Mn(2+) and Co(2+) were found to influence the binding of flavonoid to protein.     

Resolution and characterization of tryptophyl fluorescence of hen egg-white lysozyme by quenching- and time-resolved spectroscopy

The fluorescence spectral distributions of four tryptophan residues of hen egg-white lysozyme were analyzed using time-resolved and quenching-resolved fluorescence spectroscopy. Trp62 and Trp108 gave the fluorescence maxima at 352 nm and 342 nm, respectively. The fluorescence of Trp28 and Trp111 occurred only at 300-360 nm and they were observed as an unresolved emission band with a maximum and shoulder at 320 nm and 330 nm. The fluorescence quenching and decay parameters of each tryptophan residue reconfirmed that Trp62 was fully exposed to the solvent but Trp108 was sealed in the cage of the peptide chains and furthermore showed that Trp28 and Trp111 are under the influence of the larger fluctuational motion at the hydrophobic matrix box. The fluorescence responses of each tryptophan residue to the lysozyme-ligand interaction suggested that the internal fluctuation was reduced by the binding of ligand to give a distorted conformation to the hydrophobic matrix box region.     

Synthetic flavinyl peptides related to the active site of mitochondrial monoamine oxidase II. Fluorescence properties.

The fluorescence properties of various 8alpha-sulfur-linked flavinyl peptides and related flavin analogues were investigated as the pH solvent, temperature, and flavin concentration were varied. Substitution in the 8alpha position by a thioether-linked peptide brings about a marked quenching of fluorescence (up to 98% in water), a slight bathochromic shift and broadening of the fluorescence emission spectra, and a slight decrease in the fluorescence lifetimes. Oxidation of the thioether function to a sulfone partially releases this fluorescence quenching without further changes in the fluorescence emission spectra. The primary effect on the fluorescence intensity is due to an interaction between the nonbonding electrons of the thioether, the hydrogen-bonding, polar solvent, and the isoalloxazine ring. Dissolving these flavinyl peptides in nonaqueous solvents increases the fluorescence intensity as much as 20-fold. A secondary effect on flavinyl fluorescence can be attributed to a collisional quenching by the vicinal tyrosyl residue within tyrosine-containing flavinyl peptides. The fluorescence properties provide further confirmation of the identity of the synthetic and naturally obtained flavinyl peptides and of the interaction between the free-hydroxyl functions of the ribityl side chain and the thioether.     

Relative fluorescence of normal and acid lipase-deficient cultured fibroblasts following administration of pyrene decanoic acid

Skin fibroblasts, derived from normal individuals or patients with Wolman's disease (an autosomal recessive disorder due to acid lysosomal lipase deficiency) were incubated with the fluorescent fatty acid, pyrene-decanoic acid (P10). Measurements of the fluorescence intensities of the total lipid extracts indicated that equal quantities of P10 were incorporated into both cell types. The fluorescence emitted by the intact cells was subsequently recorded in a fluorescence microscope equipped with a microdetector unit, which permitted determination of the fluorescence emitted by the intact cell or by specific regions thereof. The fluorescence intensities emitted by the lipidotic cells exceeded those of their normal counterparts 2- and 5-fold when comparing the entire cells or the perinuclear region, respectively. The cells were then subjected to subcellular fractionation and an analysis of the fractions revealed that up to 85-90% of the fluorescence of the lysosome-mitochondrial pellet was derived from free pyrenedecanoic acid; the latter contributed only 15-18% to the fluorescence of the homogenate or the cytosol. There was no difference in the fluorescence of the lipid extracts from the respective fractions of the lipidotic or normal cells. However, the fluorescence emitted by the intact lysosome-mitochondrial fraction of the lipidotic cells exceeded that of its normal counterpart 2.5-fold. These data suggest that the increased fluorescence intensity of the intact lipidotic cells resulted from a higher quantum yield of free P10 molecules solubilized in the hydrophobic environment of their neutral lipid-containing storage granules.     

A fluorescence quenching test for the detection of flavonoid transformation

A novel fluorescence quenching test for the detection of flavonoid degradation by microorganisms was developed. The test is based on the ability of the flavonoids to quench the fluorescence of 1,6-diphenyl-1,3,5-hexatriene (DPH). Several members of the anthocyanidins, flavones, isoflavones, flavonols, flavanones, dihydroflavanones, chalcones, dihydrochalcones and catechins were tested with regard to their quenching properties. The anthocyanidins were the most potent quenchers of DPH fluorescence, while the flavanones, dihydroflavanones and dihydrochalcones, quenched the fluorescence only weakly. The catechins had no visible impact on DPH fluorescence. The developed test allows a quick and easy differentiation between flavonoid-degrading and flavonoid-non-degrading bacteria. The investigation of individual reactions of flavonoid transformation with the developed test system is also possible.     

Accurate determination of DNA content in single cell nuclei stained with Hoechst 33258 fluorochrome at high salt concentration

Summary In an attempt to achieve accurate quantification of DNA levels in cell nuclie, we studied the influence of salt concentration on the fluorescence of cell nuclei complexed with Hoechst-33258 (Hoe) fluorochrome. The fluorescence of cell nuclei was compared with that of extracted DNA as well as that of nucleosome core. Conformational changes in these complexes were examined by measuring both fluorescence anisotropy and fluorescence lifetime in the nanosecond region. The results showed that the fluorescence of DNA-Hoe was quenched by the nucleosomal structure, there being an associated increase in anisotropy and a decrease in the fluorescence lifetime; however, the fluorescence was restored to the orginal level by the addition of a high concentration of NaCl, CsCl, or LiCl. The reduction in fluorescence may have been due to loss of fluorescence energy caused by collision of the fluorophore with histones in the nucleosome. The addition of 1 M NaCl to the medium used for staining with Hoe greatly stabilized the fluorescence of DNA in cell nuclei. The DNA content of individual cell nuclei was determined by comparing the fluorescence of these nuclei with that of a standard DNA solution. For lymphocytes and liver ploidy cells, reasonably accurate values were obtained by applying the present method.     

共振喇曼光谱技术在恶性肿瘤诊断中的应用

目的:探讨共振喇曼光谱技术用于早期恶性肿瘤诊断的研究。方法:利用氩离子激光作为线偏振光的特点,采集偏振荧光光谱,对荧光光谱的偏振态进行分析。利用不同荧光物质的荧光可能具有不同偏振态的特点减少其它荧光物质的荧光对光谱分析的影响。血清样品产生的荧光也具有确定的偏振性。对所检测病人血清经激光分析仪进行喇曼光谱技术分析,光谱数据经计算机软件处理,自动显示图谱和数据,并直接给出各项指标及诊断提示。本结果与细胞病理学结果进行了对照研究。结果:恶性肿瘤样本176例,检测出阳性病例141例,阳性符合率为80.1%;良性肿瘤样本52例,4例阳性,假阳性率为7.7%;正常体检样本248例,检测结果均为阴性。结论:喇曼光谱技术适用于肿瘤初筛、普查及早期诊断,有推广应用前途。     

双色荧光杂交芯片在检测 P 1A 1M P I 基C 因多态性中的应用

目的：应用一种新的高通量SNP检测方法-双色荧光杂交芯片技术检测CYPIA1 MspI基因多态性。方法：收集江苏汉族人群原发性肺癌患者75例和相应对照77例,应用双色荧光杂交芯片技术检测了152例样本的CYPIAI基因MspI基因多态性,并应用PCR-RFLP技术验证双色荧光杂交芯片的特异性。结果：152例样本的CYPIAI基因双色荧光杂交芯片技术分型结果与PCR-RFLP结果完全相符,两种方法的基因型分型结果具有很好的一致性。结论：双色荧光杂交芯片技术是一个高通量SNP检测的良好工具,特异性高,在大规模人群SNP筛检中具有良好的发展前案。     

In situ and remote-sensed chlorophyll fluorescence as indicator of the physiological state of phytoplankton near the Isles Kerguelen (Southern Ocean)

Shipboard and remote-sensed Chlorophyll fluorescence were determined in the natural phytoplankton assemblage above the iron-enriched Kerguelen Plateau and the adjacent high-nutrient, low-Chlorophyll open Southern Ocean. The variance between fluorescence yield and photosynthetic efficiency was determined in combination with Chlorophyll a concentrations, irradiance and phytoplankton species distribution. A co-variance between the fluorescence measurements would allow the refinement of remote-sensing primary production algorithms. Distinct differences were found in photosynthetic efficiency and water-leaving fluorescence, with relatively high values for the Kerguelen Plateau and low values in the open ocean, reflecting the differences in Chlorophyll a concentrations. The co-variance of the fluorescence properties suggested that remote-sensed fluorescence measurements could be used to infer differences in the physiological state of the phytoplankton, hence primary production. Fluorescence yield, however, did not show the differences in the research area, most likely due to the low signal and the diurnal variation in water-leaving fluorescence.     

双歧杆菌的完整肽聚糖对小鼠腹腔巨噬细胞缝隙链接的影响

目的探讨双歧杆菌的完整肽聚糖（WPG）对巨噬细胞缝隙连接介导的细胞间通讯（GJIC）的影响。方法首先分离培养昆明小鼠腹腔巨噬细胞,然后以WPG刺激巨噬细胞,再用脂荧光探针标记细胞,最后采用激光共聚焦显微镜结合激光漂白后荧光恢复技术检测反映GJIC变化的荧光恢复程度。结果和对照组相比,以WPG刺激巨噬细胞后,其GJIC的平均荧光恢复率明显增加（P〈0.01）。结论双歧杆菌的WPG可提高巨噬细胞的缝隙连接介导的细胞间通讯。     

Deconvolution of C-phycocyanin beta-84 and beta-155 chromophore absorption and fluorescence spectra of cyanobacterium Mastigocladus laminosus.

Absorption and fluorescence spectra of the C-phycocyanin beta-subunit were quantitatively deconvoluted into component spectra of the beta-84 and beta-155 chromophores. The deconvolution procedure was based on a theoretical treatment of polarization properties. Four kinds of spectra (absorption, emission, emission polarization, and excitation polarization) measured on C-phycocyanin isolated from the cyanobacterium Mastigocladus laminosus were used as the experimental data set. Without any assumption of spectral shape, the absorption and fluorescence spectra of both chromophores were unambiguously resolved and their fluorescence quantum yields were evaluated. By combining the spectra of the alpha-subunit, independently measured, with the resolved spectra of the beta-subunit, the fluorescence and fluorescence polarization spectra and the fluorescence quantum yield of the monomer were estimated; they agree with experimental values to within an acceptable error. Further, the matrix of energy transfer rates in the monomer was estimated; it gave a significantly different result (by up to 40%) from previously estimated ones.