TEM像に迫る, リターディング法を用いた樹脂包埋生物標本のSEM反射電子観察とFIB/SEMトモグラフィーへの応用  [in Japanese] Improved Block Face Imaging and FIB/SEM Tomography from a Resin-Embedded Biological Specimen Using Back-Scattered Electrons with a Retarding Method  [in Japanese]

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Abstract

<p>重金属染色した樹脂包埋生物試料のブロック表面からのSEM反射電子組成像は超薄切片のTEM像と同等の構造情報を持つ.この組成像の像質は試料台とカラムの間に負電圧をかけるリターディング法により劇的に向上し,TEM像に迫る分解能が得られる.FIB/SEMトモグラフィー法に応用するとTEMを用いた電子線トモグラフィー法に準ずる分解能でより大きな構造を解析できる.これらの新しい生物組織観察法は,偶然得られる良い断面図に頼ることのない確実な構造観察を可能にする.</p>

<p>Recent development of scanning electron microscopy (SEM) technique, by which images are taken from back-scattered electron (BSE) from flat block surface of resin-embedded biological specimens, enabled us to obtain high contrast images similar to the ones obtained by TEM. This imaging method facilitates not only a wide area of observation but also a 3D volume analysis such as a FIB/SEM tomography method. However, the BSE image obtained by a conventional SEM under general conditions does not have sufficient contrast and resolution to observe detailed structures of the cell. Here we present a "retarding" method that drastically enhances the quality of the BSE image even when obtained by the conventional SEM. The "retarding" method reduces the primary electron energy by the negative bias voltage between the specimen and the beam column, and it provides high resolution (4 nm) and high contrast images from the resin block surface when combined with optimal SEM settings. Furthermore, when the retarding method is applied to a FIB/SEM 3D reconstruction method, it can reconstruct a larger volume than is achievable by conventional electron tomography, and its high spatial resolution permits the visualization of 3D structures of the cell such as the membrane organization of organelles.</p>

Journal

  • KENBIKYO

    KENBIKYO 47(3), 176-180, 2012-09-30

    The Japanese Society of Microscopy

References:  16

Codes

  • NII Article ID (NAID)
    10031121404
  • NII NACSIS-CAT ID (NCID)
    AA11917781
  • Text Lang
    JPN
  • Article Type
    REV
  • ISSN
    13490958
  • NDL Article ID
    024038014
  • NDL Call No.
    Z16-896
  • Data Source
    CJP  NDL  J-STAGE 
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