Localization of chromophore absorption signals in TEM with an improved prism-mirror-prism filter

被引:5
作者
Davis, JA
Heng, YM
Barfels, MMG
Bisland, SK
Gariépy, J
Ottensmeyer, FP
机构
[1] Univ Toronto, Ontario Canc Inst, Toronto, ON M5G 2M9, Canada
[2] Univ Toronto, Dept Med Biophys, Toronto, ON M5G 2M9, Canada
来源
JOURNAL OF ELECTRON MICROSCOPY | 2000年 / 49卷 / 05期
基金
英国医学研究理事会;
关键词
microanalysis; localization; chromophore; EFTEM; ESI; FITC; chlorin e(6);
D O I
10.1093/oxfordjournals.jmicro.a023852
中图分类号
TH742 [显微镜];
学科分类号
摘要
A corrected prism-mirror-prism electron energy filler with curved entrance and exit faces was designed and incorporated into a Zeiss EM902 transmission electron microscope. The installation of the new filter required little modification to the existing microscope geometry and electronics. The filter had an energy resolution of 1.1 eV over the full image plane (acceptance half angle 10 mradian). The improved energy resolution was applied in studies of the low electron energy loss region that includes molecular orbital excitations or chromophore energy absorptions. Chromophore signal behaviour under electron irradiation was characterized using embedded crystals of hematin and of the dye mercury orange. Images of these crystals confirmed the expected decrease in signal intensity on shifting the selected energy loss from the region of molecular orbital excitations (less than similar to 5 eV) to higher energy losses. Electron irradiation-induced fading of the chromophore signal from hematin and mercury orange yielded similar lie dose values of 1.1x10(5) e(-) nm(-2) and 1.4x10(5) e(-) nm(-2) respectively. In a cellular context, chromophore signals were obtained from energy-filtered images of RIF-1 cell sections containing the photosensitizer chlorin e(6) and from sections of BS-C-1 cells with cytoskeletal labelling via FITC-conjugated antibodies. The respective signals were extracted using a dose-dependent method or a shift in selected energy. Chromophore signal distributions were in agreement with fluorescence light microscopic images, but provided detail at higher spatial resolution.
引用
收藏
页码:629 / 639
页数:11
相关论文
共 27 条
[1]   Low energy loss electron microscopy of chromophores [J].
Barfels, MMG ;
Jiang, XG ;
Heng, YM ;
Arsenault, AL ;
Ottensmeyer, FP .
MICRON, 1998, 29 (2-3) :97-104
[2]  
BARFELS MMG, 1998, THESIS U TORONTO, P81
[3]  
BIHR J, 1991, P 49 ANN M EL MICR S, P354
[4]   Potentiation of chlorin e6 photodynamic activity in vitro with peptide-based intracellular vehicles [J].
Bisland, SK ;
Singh, D ;
Gariépy, J .
BIOCONJUGATE CHEMISTRY, 1999, 10 (06) :982-992
[5]  
CASTAING R, 1962, CR HEBD ACAD SCI, V255, P76
[6]   HIGH RESOLUTION ELECTRON SPECTROMETER FOR USE IN TRANSMISSION SCANNING ELECTRON MICROSCOPY [J].
CREWE, AV ;
ISAACSON, M ;
JOHNSON, D .
REVIEW OF SCIENTIFIC INSTRUMENTS, 1971, 42 (04) :411-&
[7]   CHEMICAL MEASUREMENTS OF RADIATION-DAMAGE IN ORGANIC-SAMPLES AT AND BELOW ROOM-TEMPERATURE [J].
EGERTON, RF .
ULTRAMICROSCOPY, 1980, 5 (04) :521-523
[8]  
ENGLE W, 1984, AM LAB, V16, P26
[9]   RADIATION DAMAGE IN ORGANIC CRYSTALLINE FILMS. [J].
Fryer, J.R. .
Ultramicroscopy, 1983, 14 (03) :227-236
[10]   PHOTODYNAMIC ACTIVITY OF CHLORIN-E(6) AND CHLORIN-E(6) ETHYLENEDIAMIDE INVITRO AND INVIVO [J].
GURINOVICH, GP ;
ZORINA, TE ;
MELNOV, SB ;
MELNOVA, NI ;
GURINOVICH, IF ;
GRUBINA, LA ;
SARZHEVSKAYA, MV ;
CHERENKEVICH, SN .
JOURNAL OF PHOTOCHEMISTRY AND PHOTOBIOLOGY B-BIOLOGY, 1992, 13 (01) :51-57