Inoue H, Igari T, Nishikage T, Ami K, Yoshida T, Iwai T
First Dept. of Surgery, Tokyo Medical and Dental University, Japan.
Endoscopy. 2000 Jun;32(6):439-43. doi: 10.1055/s-2000-654.
Histopathological examination for superficial gastrointestinal lesions has been mainly based upon the light microscopic examination of thin-slice specimens with hematoxylin and eosin (H&E) staining. However, it takes at least a couple of days to create a slide-glass for microscopic study. In order to obtain immediate microscopic images for untreated specimens, the authors used laser-scanning confocal microscopy (LCM) to study fresh samples of gastrointestinal mucosa.
Fresh untreated mucosal specimens from the esophagus, stomach, and colon, obtained by endoscopic pinch biopsy, polypectomy, or endoscopic mucosal resection (EMR), were fixed in normal saline and examined by LCM collecting the reflective light of a 488-nm wavelength argon laser beam. Findings from the LCM image were compared with those of conventional H&E staining in all specimens. For objective evaluation of the similarity of both pictures, the nucleus-to-cytoplasm ratio (N/C) of normal mucosa and that of cancer of the esophagus were calculated and statistically analyzed. The overall diagnostic accuracy for cancer was evaluated.
The average scanning time to obtain the LCM image of a specimen was 1.6 seconds. The LCM images acquired corresponded well to the conventional H&E light microscopic images in the esophagus, stomach, and colon. Cell wall, nucleus, cytoplasm, and tissue structural elements were simultaneously visualized by LCM scanning. A difference in N/C ratios between normal mucosa and cancer in the esophagus was statistically apparent when Welch's test (P=0.05) was applied. The overall diagnostic accuracy of the LCM study for cancer was 89.7%.
This novel method enables us to obtain an immediate serial virtual microscopic section through a fresh specimen, which has not actually been cut, although the resolution of the image obtained is still limited. These early results encourage us to develop imaging relevant to conventional histopathology alongside the development of LCM technology in the near future. We should aim at the in vivo application of LCM coupled to probes which can be introduced through the working channel of endoscopes.
浅表性胃肠道病变的组织病理学检查主要基于苏木精和伊红（H&E）染色的薄片标本的光学显微镜检查。然而，制作用于显微镜研究的载玻片至少需要几天时间。为了获取未经处理标本的即时显微镜图像，作者使用激光扫描共聚焦显微镜（LCM）研究胃肠道黏膜新鲜样本。
通过内镜钳取活检、息肉切除术或内镜黏膜切除术（EMR）获得的食管、胃和结肠的新鲜未经处理的黏膜标本，用生理盐水固定，通过LCM收集488纳米波长氩激光束的反射光进行检查。将LCM图像的结果与所有标本中传统H&E染色的结果进行比较。为了客观评估两张图片的相似性，计算并统计分析正常黏膜和食管癌的核质比（N/C）。评估癌症的总体诊断准确性。
获取标本LCM图像的平均扫描时间为1.6秒。在食管、胃和结肠中，获取的LCM图像与传统H&E光学显微镜图像非常吻合。通过LCM扫描可同时观察到细胞壁、细胞核、细胞质和组织结构成分。应用韦尔奇检验（P = 0.05）时，食管正常黏膜和癌组织的N/C比值差异具有统计学意义。LCM研究对癌症的总体诊断准确性为89.7%。
这种新方法使我们能够通过新鲜标本获得即时的连续虚拟显微切片，尽管所获得图像的分辨率仍然有限，但该标本实际上并未被切割。这些早期结果鼓励我们在不久的将来随着LCM技术的发展，开发与传统组织病理学相关的成像技术。我们的目标应该是将LCM与可通过内镜工作通道引入的探针结合应用于体内。
