Background: Using a combination of autofluorescence from cells and second-harmonic generation (SHG) signal from collagen, multiphoton microscopy (MPM) imaging can provide detailed real-time information on tissue architecture and cellular morphology in live tissue without administration of exogenous contrast agents. The purpose of this study is to evaluate the feasibility of using MPM to histologically diagnose gastric cancer by using fresh, unfixed, unstained gastric specimens, compared with gold-standard hematoxylin-eosin (H-E)-stained histopathology.
Methods: A pilot study was performed between June 2009 and December 2009. Ten cases with gastric carcinoma confirmed by preoperative endoscopic biopsy underwent radical gastrectomy. The fresh specimen was opened, and a piece of cancer tissue and a piece of normal tissue each with a size of 1-1.5 cm across and 0.2 cm in thickness were taken and snap-frozen. A 5-μm slide was sectioned for MPM examination, and the remainder of the tissue went through routine histopathological procedure. MPM images and H-E-stained images were compared by the same attending pathologist.
Results: MPM images were acquired by two channels: broadband autofluorescence from cells, and SHG from tissue collagen. Peak multiphoton autofluorescence intensity was detected in mucosa excited at 800 nm. Cancer cells, characterized by irregular size and shape, enlarged nuclei, and increased nuclear-to-cytoplasmic ratio, were identified by MPM images, which were confirmed by H-E-stained images. Regular architectures of gastric pits and gastric glands in the normal tissue of the same specimens were clearly revealed by MPM images, which were comparable to H-E-stained images.
Conclusions: It is feasible to use MPM to diagnose gastric cancer by "optical biopsy." With miniaturization and integration of endoscopy, MPM has the potential to provide real-time histological diagnosis without invasive biopsy for gastric cancer in the future.