Quantitative chemical analysis of ocular melanosomes in stained and non-stained tissues

Energy-filtered Analytical Electron Microscopy (AEM) was used to image the ultrastructure and determine quantitatively the chemical composition of rat melanosomes of the choroid and the Retinal Pigment Epithelium (RPE). For the first time, the effect of staining in elemental analysis of melanosomes was investigated. Detection limits and accuracies of the applied methods were determined. Compared to previous work applying only quantitative Energy Dispersive X-ray microanalysis (EDX) in the TEM (Eibl, O., et al., 2006. Micron 37, 262), here we present a combined quantitative EDX and Electron Energy Loss Spectroscopy (EELS) analysis, including N. This yields the fraction of eumelanin and pheomelanin in melanosomes by the S/N mole fraction ratio. Melanosomes of the sepia ink sac, used as eumelanin standard, showed an S/N mole fraction ratio of <0.004. Thus, they consist primarily of eumelanin as reported by degradation analysis. In contrast, melanosomes of the rats contained mixed melanin with significant amounts of pheomelanin (S/N 0.02) in the RPE and the choroid. Consistent with the previous publication, it was shown that oxygen mole fractions are especially large in melanosomes (7-10 at.%) compared to other cell compartments, e.g. 2-4 at.% oxygen in the cytoplasm. In the melanosomes of non-stained tissue, the oxygen mole fraction clearly correlated with the Ca mole fraction. EDX spectra used for quantitative analysis had about 15,000 net counts under the oxygen peak, which is necessary to obtain (i) a small statistical error for oxygen and (ii) optimum minimum detectable mole fractions for S, Ca and transition metals. The precise determination of the oxygen mole fraction in melanosomes is important for understanding metabolism. Therefore, a detailed analysis was carried out on the possible errors affecting quantification. While O, S, and N mole fractions yielded similar results in stained and non-stained ocular melanosomes of rats, transition metals can only be determined reliably in non-stained tissues. High-precision EDX analysis of melanosomes yielded minimum detectable mole fractions of less than 0.04 at.% for Cu and Zn, these elements were present in melanosomes with mole fractions of about 0.3 at.% and 0.1at.%, respectively. Zn is of great importance for metabolism and for age related macular degeneration. Its mole fraction in melanosomes of rats is large enough to be detected and to be quantitatively analyzed by EDX spectroscopy. Ultrastructural information can now be correlated to the elemental composition. This is important to better understand the physical and chemical properties of melanosomal metabolism and turnover.     

Analysis of temperature and 1 MeV proton irradiation effects on the light emission in bulk silicon (npn) emitter-base bipolar junctions

In this work, we present the temperature and 1 MeV irradiation proton effects on the light emission in bulk silicon emitter-base junctions for direct and reverse polarizations. Our samples were exposed at room temperature to 5.3 × 10⁸, 5.3 × 10¹⁰, 5 × 10¹¹, 5 × 10¹² and 5 × 10¹³ p cm⁻². The spectral range for which electroluminescence spectrums were recorded for forward and reverse polarizations is 0.6–2 eV. For forward bias, EL maximum intensity occurs at 1.0923 ± 0.0001 eV (structure (a)) which decreases as function of irradiation fluencies. For reverse bias, the spectra contain two structures (b) and (c). The first structure (c) occurred at 1.6243 ± 0.0013 eV is independent of irradiation while the second structure (b) decreases as function of fluencies irradiation. The Gaussian deconvolution of (b) shows two sub-structures (b₁) and (b₂) which are located, respectively, at 0.8064 ± 0.0004 eV and 0.9917 ± 0.0016 eV. We studied temperature dependence of full width at half-maximum (FWHM) and we found that the phonons involved in (a), (b₁) and (b₂) on the one hand and (c) on the other hand are not the same. Moreover, we obtained from the study of EL intensity temperature dependence that the activation energies of (a), (b₁) and (b₂) are identical and differ from that of (c). These effects enable us to conclude that visible light emission does not have the same origin as that in infra-red. From these observations, we can attribute the structures (a) and (b) to indirect inter-bands transitions and (c) to a direct intra-band transitions.     

Effect of Melanomal Proteins on Sepia Melanin Assembly

Melanins are phenol-based pigments with the potential for widespread applications, including bioelectronics and tissue engineering. The concentration-dependent structural transition of sepia melanin in water is analyzed. This biopolymer at high concentration gives the well-known nanospheres, whereas sample dilution gives unforeseen nanofibres exhibiting the structural features of mature amyloid fibrils. We propose a mechanism of pigment self-assembly dependent on the interaction of residual melanosomal protein(s) with eumelanin heteropolymer. Our results contribute to understanding the peculiar physicochemical properties of this ubiquitous pigment.     

Ultrastructural changes in the melanocytes of aging human choroid

Retinal pigment epithelial cells as well as choroidal melanocytes (CM) possess melanin granules. The former show clear, age-related changes (formation of lipofuscin granules with a concomitant decrease in melanin content); however, data on changes in the CM with aging are fairly limited. We examined CM in human macular and mid-peripheral areas by light- and transmission electron microscopy in 50–94 year-old donor eyes (N = 12). Unlike in the choroid of lower ages, the melanocytes from aging choroid (>75 years) showed partial fusion of about 8–15 melanosomes, forming rosettes-like structures. Besides, there was evidence of emptiness in cytoplasm caused by the loss of melanosomes in aged CM, as was confirmed by quantification in macular part of choroid. In advanced aged eyes (85–94-year-old), the CM possessed many lipid droplets as well as irregular lipofuscin granules, the latter had a tendency to fuse with melanosomes, as happens in aged retinal pigment epithelium. Macrophages in their cytoplasm contained abundant irregular as well as clumped melanosomes of variable size, suggesting that damaged granules/melanocytes are cleared by these phagocytes. These obvious changes in the CM are likely to make the choroid prone to damage by visible light.     

Optical constants of Tl4Ga3InSe8 layered single crystals

The optical properties of Tl₄Ga₃InSe₈ layered single crystals have been studied by means of transmission and reflection measurements in the wavelength range of 500–1100 nm. The analysis of the room temperature absorption data revealed the presence of both optical indirect and direct transitions with band gap energies of 1.94 and 2.20 eV, respectively. Transmission measurements carried out in the temperature range of 10–300 K revealed that the rate of change of the indirect band gap with temperature is γ=−4.1×10⁻⁴ eV/K. The absolute zero value of the band gap energy was obtained as E_gi(0)=2.03 eV. The dispersion of the refractive index is discussed in terms of the Wemple–DiDomenico single-effective-oscillator model. The refractive index dispersion parameters: oscillator energy, dispersion energy, oscillator strength and zero-frequency refractive index were found to be 4.10 eV, 23.17 eV, 6.21×10¹³ m⁻² and 2.58, respectively. From X-ray powder diffraction study, the parameters of monoclinic unit cell were determined.     

Optical properties and electronic transitions of SnO2 thin films by reflection electron energy loss spectroscopy

Optical properties of SnO₂ thin films in the 4–60 eV energy range are determined by reflection electron energy loss spectroscopy. Bulk and surface electron loss functions, real and imaginary parts of the dielectric function, refraction index, extinction and absorption coefficients are obtained from the analysis of the electron energy loss spectra. Electronic transitions are identified through the interpretation of the optical data. The samples (250–500 nm thick) were produced by ion beam-induced chemical vapor deposition. It is found that the compacity of the SnO₂ thin films affects their optical properties and therefore the relative intensity of the observed electronic transitions. The advantages of this method to determine optical properties of thin films are discussed. Inelastic mean free paths (6.2, 17 and 41 Å for electrons traveling in SnO₂ with kinetic energies of 300, 800 and 2000 eV, respectively) are obtained from the corresponding inelastic electron scattering cross-sections.     

Ti/Al p-GaN Schottky barrier height determined by C–V measurements

Data are presented on the rigorous method of capacitance–voltage (C–V) measurements to the barrier height of Ti/Al p-GaN Schottky junction. For a sample with Hall concentration of 5.5 × 10¹⁶/cm³ the upper limit of the modulation frequency leading the full response of the activated carriers is defined as 1.5 kHz from the capacitance versus modulation frequency (C–f) plot. The activation energy of the Mg acceptors determined from the temperature-dependent C–f plot is 0.12 eV. The barrier height estimated with this activation energy and the intercept voltage of the 1/C²–V plot drawn with the 1.5 kHz C–V data is 1.43 eV at 300 K and 1.41 eV at 500 K. This is the most reliable barrier height ever reported. A reliable room temperature C–V doping profile is demonstrated using the 1.5 KHz modulation, which is sensitive enough to resolve the presence of a 15 nm thin highly doped (8 × 10¹⁸/cm³) layer formed near the surface.     

Surface diffusion of dysprosium on the W(1 1 1) facet

Diffusion of dysprosium on the (1 1 1) facet of a tungsten micromonocrystal was investigated by means of spectral analysis of field emission current fluctuations. The experimental spectral density functions of the current fluctuations were analysed by using Gesley and Swanson’s theoretical spectral density function, which enables to determine the surface diffusion coefficient D for dysprosium. Derived from the temperature dependence of D, the diffusion activation energy E is presented for some Dy coverages θ_(1 1 1). In the temperature range 400–600 K, the E first drops from 1.25 eV per atom at θ₍₁₁₁₎≈0.25 ML to 0.48 eV per atom at θ₍₁₁₁₎≈1 ML (corresponding to the minimum of the work function of the system), then increases to 1.03 eV per atom at θ₍₁₁₁₎≈1.3 ML. The results are discussed from the aspects of the substrate structure and interaction in the adsorbed layer.     

A metastable de-excitation spectroscopy (MDS) study on oxygen adsorption on a polycrystalline zirconium surface

The adsorption of oxygen on a polycrystalline zirconium surface at room temperature has been studied by metastable de-excitation spectroscopy (MDS) in conjunction with UPS and AES. From the analysis of the measured spectra, we have shown the following. (1) At the initial stage of oxygen adsorption (exposure <1.2 L), the surface density of states (SDOS) of zirconium changes little at around the Fermi level (E_F), while it decreases appreciably at 1–2 eV below E_F (E_B=1–2 eV) by oxygen adsorption. (2) The SDOS at E_B=0–2 eV decreases with increasing oxygen exposure at >1.2 L and disappears at >8 L. (3) The oxygen 2p states (E_B=5–8 eV) are localized at the subsurface region at oxygen exposure 0–2 L. (4) The ZrO₂ phase appears at the outermost zirconium surface at around 2 L, then grows with increasing exposure, and finally dominates at >8 L. It is suggested that two different phases (ZrO₂ phase and that in which oxygen occupies subsurface sites) coexist at the outermost surface at 2–8 L.     

Investigation of the role of oxygen in NO reduction by C2H4 on the surface of stepped Pt(3 3 2)

The influence of pre-dosed oxygen on NO–C₂H₄ interactions on the surface of stepped Pt(3 3 2) has been investigated using Fourier transform infrared reflection–absorption spectroscopy (FTIR-RAS) and thermal desorption spectroscopy (TDS). The presence of oxygen significantly suppresses the adsorption of NO on the steps of Pt(3 3 2), leading to a very specific adsorption state for NO molecules when oxygen–NO co-adlayers are annealed to 350 K (assigned as atop NO on step edges). An oxygen-exchange reaction also takes place between these two kinds of adsorbed molecules, but there appears to be no other chemical reaction, which can result in the formation of higher-valence NOx.

C₂H₄ molecules which are post-dosed at 250 K to adlayers consisting of ¹⁸O and NO do not have strong interactions with either the NO or the ¹⁸O atoms. In particular, interactions which may result in the formation of new surface species that are intermediates for N₂ production appear to be absent. However, C₂H₄ is oxidized to C¹⁸O₂ by ¹⁸O atoms at higher annealing temperature. This reaction scavenges surface ¹⁸O atoms quickly, and the adsorption of NO molecules on step sites is therefore quickly restored. As a consequence, NO dissociation on steps proceeds very effectively, giving rise to N₂ desorption which closely resembles that following only NO exposure on a clean Pt(3 3 2), both in peak intensity and desorption temperature. It is concluded that the presence of ¹⁸O₂ in the selective catalytic reduction (SCR) of NO with C₂H₄ on the surface of Pt(3 3 2) does not play a role of activating reactants.     

Cathodoluminescence properties of silicon nanocrystallites embedded in silicon oxide thin films

Cathodoluminescence (CL) spectra for the Si nanocrystallites embedded in a matrix of silicon oxide films are measured at room temperature. The CL spectra consist of two principal bands whose peak energies are in a near-infrared (NIR) region (<1.6 eV) and in a blue region (2.6 eV), respectively. The spectral feature of the NIR CL band is similar to the corresponding PL spectra. The strong correlation between the presence of Si nanocrystallites and the formation of the NIR CL band are found as well as the PL spectrum. The peak energy of the blue CL band is slightly lower than that of the luminescence band originating from oxygen vacancies (≡Si–Si≡) in SiO₂. Therefore, the blue CL band is considered to come from Si_n clusters with n3 in the oxide matrix. Under irradiation of electron beams, degradation of the intensity is observed for both the CL bands but the decay characteristics are different.     

Optical characterization of amorphous Ga50Se45S5 films

The transmission spectra of thermally evaporated Ga₅₀Se₄₅S₅ films were measured over the wavelength range 300–900 nm. A simple method, suggested by Swanepoel, was used for the determination of the optical constants and thickness of the films. Increasing the thickness of the film beyond 450 nm does not affect the optical constants. The dependence of the absorption coefficient on the photon energy (hν) at the edge of the absorption band is well described by the relation hν=β(hν−E_opt)² with an optical gap equals 2.4 eV. A good fit of the experimental points with Tauc relation indicates that non-direct transition is the most probable mechanism responsible for the photon absorption inside the investigated film.     

Surface gap and surface electronic states in CuGeO3 single crystal

The surface and bulk electronic excitations of CuGeO₃ are investigated by means of electron energy loss and polarized X-ray absorption spectroscopy. CuGeO₃ shows a surface charge transfer gap of about 3.0±0.3 eV. The unoccupied oxygen derived density of states, as probed by X-ray absorption at the O 1s edge, is in good agreement with recent many-body calculations.     

Adsorption and dissociation of CO on Fe(1 1 0) from first principles

We employ spin-polarized periodic density functional theory (DFT) to characterize CO adsorption and dissociation on the Fe(1 1 0) surface. We investigate the site preference for CO on Fe(1 1 0) at θ_CO = 0.25 and 0.5 monolayer (ML), for different functional forms of the generalized gradient approximation (GGA) to electron exchange and correlation within DFT. At 0.25 ML, we predict the existence of a new ordered structure comparable in stability to one proposed previously. At 0.5 ML, we confirm the preference of a distorted on-top adsorption configuration suggested by experiment. The calculated heats of adsorption, CO stretching frequencies, and work function changes agree well with experiment. When dissociating from the on-top site, we predict that CO first moves off the on-top site and then goes through a lying-down transition state with a barrier of 1.52 eV. Diffusion of CO on Fe(1 1 0) from the on-top site to the long-bridge site is predicted to have a very small barrier of 0.1 eV. Dissociation of CO from the long-bridge site goes through the same transition state as from the on-top site, but the former has a slightly lower barrier. After dissociation, O atoms remain on the surface while C atoms are embedded into Fe(1 1 0), indicating C atoms may readily diffuse into Fe(1 1 0).     

Haliotis tuberculata hemocyanin (HtH): analysis of oligomeric stability of HtH1 and HtH2, and comparison with keyhole limpet hemocyanin KLH1 and KLH2

The multimeric/higher oligomeric states of the two isoforms of Haliotis tuberculata hemocyanin (HtH1 and HtH2) have been assessed by transmission electron microscopy (TEM) of negatively stained specimens, for comparison with previously published structural data from keyhole limpet hemocyanin (KLH1 and KLH2) [see Harris, J.R., Gebauer, W., Guderian, F.U., Markl, J., 1997a. Keyhole limpet hemocyanin (KLH), I: Reassociation from Immucothel followed by separation of KLH1 and KLH2. Micron, 28, 31–41; Harris, J.R., Gebauer, W., Söhngen, S.M., Nermut, M.V., Markl, J., 1997b. Keyhole limpet hemocyanin (KLH). II: Characteristic reassociation properties of purified KLH1 and KLH2. Micron, 28, 43–56; Harris, J.R., Gebauer, W., Adrian, M., Markl, J., 1998. Keyhole limpet hemocyanin (KLH): Slow in vitro reassociation of KLH1 and KLH2 from Immucothel. Micron, 29, 329–339]. In purified samples of both HtH isoforms, the hollow cylindrical ca 8 MDa didecamer predominates together with a small number of decamers, but tri- and longer multidecamers are detectable only in the HtH2. The stability of the two HtH isoforms under varying ionic conditions have been monitored, thereby enabling conditions for the production of stable decamers to be established. The ability of these decamers to reform multimers in the presence of 10 and 100 mM concentrations of calcium and magnesium ions in Tris–HCl buffer (pH 7.4), and also of individual HtH1 and HtH2 subunits (produced by pH 9.6 dissociation in glycine-NaOH buffer), to reassociate in the presence of calcium and magnesium ions, has been assessed. For the HtH1 decamers, the predominant multimeric product is the didecamer at 10 and 100 mM calcium and magnesium concentrations, whereas for the HtH2 decamers, large numbers of multidecamers are produced, with the reaction proceeding more completely at the higher calcium and magnesium concentration. With the HtH1 subunit, reassociation in the presence of 10 and 100 mM calcium and magnesium ions yielded an almost 100% conversion into didecamers, whereas the HtH2 subunit produced a mixture containing large numbers of short multidecamers and relatively few didecamers, together with a considerable number of smaller diameter helical/tubular polymers. The association properties of the HtH1 and HtH2 decamers, and the subunit reassociation, firmly indicate the integrity and structural competency of the protein under the experimental conditions used. Data on the association of KLH2 decamers is also presented, which together with previously published data on the association KLH1 decamers and the reassociation of KLH1 and KLH2 subunits, enables a detailed comparison of the two hemocyanin isoforms from both molluscan species to be made. Biochemical manipulation of the oligomer states and the subunit reassociation of molluscan hemocyanins can usefully be assessed by the study of negatively stained TEM specimens.     

Adsorption of oxygen on Au(111) by exposure to ozone

Atomic oxygen coverages of up to 1.2 ML may be cleanly adsorbed on the Au(111) surface by exposure to O₃ at 300 K. We have studied the adsorbed oxygen layer by AES, XPS, HREELS, LEED, work function measurements and TPD. A plot of the O(519 eV)/Au(239 eV) AES ratio versus coverage is nearly linear, but a small change in slope occurs at Θ_O=0.9 ML. LEED observations show no ordered superlattice for the oxygen overlayer for any coverage studied. One-dimensional ordering of the adlayer occurs at low coverages, and disordering of the substrate occurs at higher coverages. Adsorption of 1.0 ML of oxygen on Au(111) increases the work function by +0.80 eV, indicating electron transfer from the Au substrate into an oxygen adlayer. The O(1s) peak in XPS has a binding energy of 530.1 eV, showing only a small (0.3 eV) shift to a higher binding energy with increasing oxygen coverage. No shift was detected for the Au 4f_7/2 peak due to adsorption. All oxygen is removed by thermal desorption of O₂ to leave a clean Au(111) surface after heating to 600 K. TPD spectra initially show an O₂ desorption peak at 520 K at low Θ_O, and the peak shifts to higher temperatures for increasing oxygen coverages up to Θ_O=0.22 ML. Above this coverage, the peak shifts very slightly to higher temperatures, resulting in a peak at 550 K at Θ_O=1.2 ML. Analysis of the TPD data indicates that the desorption of O₂ from Au(111) can be described by first-order kinetics with an activation energy for O₂ desorption of 30 kcal mol⁻¹ near saturation coverage. We estimate a value for the Au–O bond dissociation energy D(Au–O) to be 56 kcal mol⁻¹.     

The electrical characterization of Zn(Phen)q/p-type Si/Al diode with interfacial layer by current–voltage characteristics

The current–voltage characteristics of Zinc (II) [(8-hydroxyquinoline)(1,10-phenanthroline)] complex (Zn(phen)q)/p-type Si/Al diode with interfacial layer have been investigated. The barrier height and ideality factor of the diode were found to be 0.71 eV and 2.05. Zn(phen)q/p-type Si/Al diode shows a metal–insulator–semiconductor structure resulted from presence of series resistance and an interfacial layer. The n and φ_B values obtained the presence of interfacial layer are 1.02 and 0.70 eV, respectively. The effect of series resistance was evaluated using a method developed by Cheung. The R_s and n values were determined from the d ln(I)/dV plot and were found to be 30.43 kΩ and 2.16, respectively. The barrier height and R_s values were calculated from the H(I)–I plot and were found to be 0.70 eV and 30.99 kΩ. The density of the interface states of the Zn(phen)q/p-type Si/Al diode was calculated and was found to be an order of 10¹³ eV⁻¹ cm⁻².     

Ion-induced secondary electron and negative ion emission from Mo/O

Absolute yields of secondary electrons and negative ions resulting from collisions of Na⁺ with Mo(100) and a polycrystalline molybdenum surface have been measured as a function of the oxygen coverage of the surface for impact energies below 500 eV. The sputtered negative ions have been identified with mass spectroscopy, and O⁻ is found to be the dominant sputtered negative ion for the surfaces at all oxygen coverages and impact energies. Both the electron and O⁻ yields have an impact energy threshold at about 50 eV and exhibit a strong dependence on oxygen coverage. The kinetic energy distributions of the secondary electrons and sputtered O⁻ were determined as functions of the oxygen coverage and impact energy. The distributions for O⁻ are characterized by a narrow low-energy peak (at 1–2 eV) followed by a low-level high-energy tail. The secondary electrons have a narrow (FWHM 1–2 eV) kinetic energy distribution, centered approximately at 1–2 eV. The shapes of the distributions and their most probable energies are essentially invariant with impact energy, oxygen coverage and the nature of the Mo surface. The emission is explained and analyzed in terms of a simple model which involves a collision-induced electronic excitation of the MoO⁻ surface state. The decay of this excited state leads to the production of both secondary electrons and O⁻ with energy distributions and yields comparable to those observed.     

Electrical conductivity, optical and metal–semiconductor contact properties of organic semiconductor based on MEH-PPV/fullerene blend

The electrical conductivity, optical and metal–semiconductor contact properties of the MEH-PPV:C₇₀ organic semiconductor have been investigated. The electrical conductivity results show that the MEH-PPV:C₇₀ film is an organic semiconductor. The optical band gap of the film was found to be 2.06 eV and the fundamental absorption edge in the film is formed by the direct allowed transitions. The refractive index dispersion curve of the film obeys the single oscillator model and E_d and E_o dispersion parameters were found to be 10.61 and 3.89 eV, respectively. The electrical characterization of the ITO/MEH-PPV:C₇₀ diode have been investigated by current–voltage characteristics. ITO/MEH-PPV:C₇₀ diode indicates a non-ideal current–voltage behavior with ideality factor n (2.50) and barrier height φ_B (0.90 eV) values.     

Fe掺杂GaN光电特性的第一性原理研究

基于密度泛函理论体系,计算了本征GaN材料和12.5%的Fe掺杂GaN体系的光电特性,分析了晶体结构、能带结构和电子态分布、介电函数、吸收系数、折射率、反射率、能量损失谱和消光系数,从理论上讨论了掺杂对体系光电特性的影响.计算所得理想GaN的禁带宽度为3.41 eV,Fe的重掺杂体系明显变窄,为3.06 eV,但仍为直接带隙半导体.本征GaN材料与Fe掺杂GaN体系的静态介电常数为5.74和6.20,折射率为2.39和2.48,能量损失最大值在20.02 eV和18.96 eV,最大吸收系数能量均在13.80 eV左右.计算结果为Fe掺杂GaN高压光电导开关材料及器件的进一步研究提供了有力的理论依据和实验支持.