Intraoperative 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.

中文翻译：

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用于检测前列腺癌的非线性显微术：根治性前列腺切除术的敏感性和特异性分析。

使用冰冻切片在根治性前列腺切除术期间对标本进行术中评估可能是时间和劳动密集型的。非线性显微镜 (NLM) 是一种荧光显微镜技术，可以在新鲜切除的组织中快速生成非常类似于 H&E 组织学的图像，而无需冷冻或切片机切片。标本用细胞核和细胞质/基质荧光团染色，NLM 评估可以在总计 3 分钟内开始。荧光信号可以使用 H&E 色标显示，便于病理学家解释。本研究与福尔马林固定、石蜡包埋的 H&E 组织学金标准相比，评估了 NLM 图像盲读中前列腺癌检测的准确性。从 40 名接受根治性前列腺切除术的患者中获得了总共 122 个新鲜切除的前列腺标本。前列腺被粗略地解剖成约 10 × 10 毫米、厚度为 1-4 毫米的标本，染色 2 分钟用于细胞核和细胞质/基质对比，然后用盐水冲洗 30 秒。获取 NLM 图像并将多个图像拼接在一起以生成适合阅读的大视野、厘米级数字图像。然后对标本进行标准石蜡 H&E 处理。研究方案包括训练、预测试和盲读阶段。清洗期后，病理学家读取相应的石蜡 H&E 载玻片。与石蜡 H&E 相比，三位病理学家在 NLM 上检测癌症的灵敏度达到 95% 或更高，特异性为 100%。汇总的敏感性和特异性分别为 97.3%（93.7-99.1%；95% 置信区间）和 100.0% (97.0-100.0%)。NLM 阅读的观察者间协议具有 Fleiss κ = 0.95。高癌症检测准确性和快速标本制备表明 NLM 可用于根治性前列腺切除术的术中评估。