A novel method of virtual histopathology using laser-scanning confocal microscopy in-vitro with untreated fresh specimens from the gastrointestinal mucosa

Background and Study Aims: Histopathological examination for superficial ga strointestinal lesions has been mainly based upon the light microscopic exa mination 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 mi croscopic study. In order to obtain immediate microscopic images for untrea ted 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 examine d 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 wa s 1.6 seconds. The LCM images acquired corresponded well to the conventiona l H&E light microscopic images in the esophagns, stomach, and colon. Cell w all, nucleus, cytoplasm, and tissue structural elements were simultaneously visualized by LCM scanning. A difference in NIC ratios between normal muco sa 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 vir tual microscopic section through a fresh specimen, which has not actually b een cut, although the resolution of the image obtained is still limited. Th ese early results encourage us to develop imaging relevant to conventional histopathology alongside the development of LCM technology in the near futu re, We should aim at the in vivo application of LCM coupled to probes which can be introduced through the working channel of endoscopes.