A novel method of virtual histopathology using laser-scanning confocal microscopy in-vitro with untreated fresh specimens from the gastrointestinal mucosa
A novel method of virtual histopathology using laser-scanning confocal microscopy in-vitro with untreated fresh specimens from the gastrointestinal mucosa
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DOI：
10.1055/s-2000-654
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发表时间：
2000-06-01
期刊：
影响因子：
9.3
通讯作者：
Iwai, T
中科院分区：
文献类型：
--
作者：
Inoue, H;Igari, T;Iwai, T
背景和研究目的：浅表性胃肠道病变的组织学检查主要基于苏木精和伊红（H&E）染色的薄片标本的光镜检查。然而，至少需要几天的时间来创建一个用于显微镜研究的载玻片。为了获得未经处理的标本的即时显微图像，作者使用激光扫描共聚焦显微镜（LCM）研究胃肠道粘膜的新鲜样本。材料和方法：通过内镜夹捏活检、息肉切除术或内镜粘膜切除术（EMR）获得的来自食管、胃和结肠的新鲜未经处理的粘膜标本，固定在生理盐水中，并通过LCM收集488 nm波长氩激光束的反射光进行检查。在所有标本中，将LCM图像的结果与常规H&E染色的结果进行比较。为了客观评价两种图像的相似性，计算正常粘膜和食管癌的核质比（N/C）并进行统计学分析。结果：获得标本LCM图像的平均扫描时间为1.6秒。所获得的LCM图像与食管、胃和结肠中的常规H&E光学显微镜图像对应良好。LCM扫描同时显示细胞壁、细胞核、细胞质和组织结构成分。当应用Welch检验（P=0.05）时，正常粘膜和食管癌之间NIC比率的差异在统计学上是明显的。LCM研究癌症的总体诊断准确率为89.7%，结论：这种新方法使我们能够通过新鲜标本获得即时的系列虚拟显微镜切片，该标本实际上尚未切割，尽管所获得的图像分辨率仍然有限。这些早期的结果鼓励我们在不久的将来开发与传统组织病理学相关的成像以及LCM技术的发展，我们的目标应该是将LCM耦合到可以通过内窥镜的工作通道引入的探针的体内应用。
Background and Study Aims: 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,Materials and Methods: 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.Results: 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 esophagns, stomach, and colon. Cell wall, nucleus, cytoplasm, and tissue structural elements were simultaneously visualized by LCM scanning. A difference in NIC 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 %,Conclusions: 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.