Ponchiardi, CeciliaRosen, SeymourCahill, LucasWu, YuboYoshitake, TadayukiGiacomelli, MichaelWagner, AndrewFujimoto, James2022-08-052019-11-19Ponchiardi, Cecilia, Seymour Rosen, Lucas Cahill, Yubo Wu, Tadayuki Yoshitake, Michael Giacomelli, Andrew Wagner et al. "Nonlinear microscopy for detection of prostate cancer: analysis of sensitivity and specificity in radical prostatectomies." Modern Pathology 33, no. 5 (2019): 916-923. DOI: 10.1038/s41379-019-0408-40893-39521530-0285https://nrs.harvard.edu/URN-3:HUL.INSTREPOS:37372943Intraoperative evaluation of specimens during radical prostatectomy using frozen sections can be time and labor intensive. Nonlinear microscopy (NLM) is a fluorescence microscopy technique that can rapidly generate images that closely resemble H&E histology in freshly excised tissue, without requiring freezing or microtome sectioning. Specimens are stained with nuclear and cytoplasmic/stromal fluorophores, and NLM evaluation can begin within 3 min of grossing. Fluorescence signals can be displayed using an H&E color scale, facilitating pathologist interpretation. This study evaluates the accuracy of prostate cancer detection in a blinded reading of NLM images compared with the gold standard of formalin-fixed, paraffin-embedded H&E histology. A total of 122 freshly excised prostate specimens were obtained from 40 patients undergoing radical prostatectomy. The prostates were grossed, dissected into specimens of ~10 × 10 mm with 1–4 mm thickness, stained for 2 min for nuclear and cytoplasmic/stromal contrast, and then rinsed with saline for 30 s. NLM images were acquired and multiple images were stitched together to generate large field of view, centimeter-scale digital images suitable for reading. Specimens were then processed for standard paraffin H&E. The study protocol consisted of training, pretesting, and blinded reading phases. After a washout period, pathologists read corresponding paraffin H&E slides. Three pathologists achieved a 95% or greater sensitivity with 100% specificity for detecting cancer on NLM compared with paraffin H&E. Pooled sensitivity and specificity was 97.3% (93.7–99.1%; 95% confidence interval) and 100.0% (97.0–100.0%), respectively. Interobserver agreement for NLM reading had a Fleiss κ = 0.95. The high cancer detection accuracy and rapid specimen preparation suggest that NLM may be useful for intraoperative evaluation in radical prostatectomy.en-USPathology and Forensic MedicineJournal Article2022-08-0510.1038/s41379-019-0408-4