Label-free intraoperative photoacoustic microscopy for rapid diagnosis of tissue biopsies with specimen integrity

摘要（30行） 术中通常需要冰冻切片诊断来指导手术。虽然这种方法 尽管它仍然是不可替代的，但它存在一系列局限性：组织损失，冷冻伪影，脂肪的次优切割， 标本，无法诊断骨病变，延长周转时间。 目的是开发一种多通道亚细胞分辨光声显微镜系统， 解决所有上述局限性，并为病理学家提供诊断图像， 或者与现有技术相同，具有匹配或超过标准透射光的放大率 显微镜，并且不需要病理学家进行额外的诊断培训。 为达致这个目标，我们会致力达致以下两个具体目标： 具体目标1：开发多通道亚细胞分辨率PAM系统，用于快速分析和诊断 组织活检 A.多通道亚细胞分辨PAM系统的研制 B。开发定制的组织活检保持器，用于快速组织活检处理和PAM成像 具体目标2：优化和验证多通道亚细胞分辨率PAM， 准确、高效的术中器械 A.优化图像采集，以最大限度地区分细胞核、细胞质和间质 具有最小组织降解的性能 B。优化采集后算法以生成类似于经典苏木素的伪彩色图像 和伊红（H&E）切片，叠加独特的成像模式（水肿和胶原蛋白） C.通过与H&E切片进行比较，验证PAM生成的图像用于诊断目的的利用率， 其通过福尔马林固定的石蜡包埋（当前的金标准）制备 D.通过比较PAM生成的图像与 H&E切片，由冷冻切片制备（目前的术中金标准） 影响：所提出的技术将通过解决当前技术的所有局限性来代表一个重大突破。 基于冰冻切片的诊断，并在术中提供具有H&E等效质量的图像。旨在 为手术活检引入快速和节省组织的诊断方法，对医疗保健产生积极影响 系统和病人护理。减少冻结切片解释的周转时间将大大 减少在手术室花费的时间，从而降低患者风险和成本。的能力 评估没有组织损失的小样本的适当性将减少重新活检程序的需要。 虚拟生成的图像可以远程解释，以便较小的医院可以从程序中受益 目前只提供给较大的三级医院和大学中心。

ABSTRACT (30 Lines) Frozen section diagnoses are typically requested intraoperatively to guide the surgery. While this methodology is still irreplaceable, it suffers from a series of limitations: tissue loss, freezing artifacts, suboptimal cutting of fatty specimens, inability to diagnose bony lesions, and prolonged turnaround time. The aim is to develop a multi-channel subcellular resolution photoacoustic microscopy (PAM) system capable of addressing all the aforementioned limitations and providing pathologists with diagnostic images comparable or identical to the state of the art, with magnification matching or exceeding that of standard transmission light microscopes, and requiring no additional training for pathologists to diagnose. To achieve our objective, we will pursue the following two specific aims: Specific Aim 1: Develop a multi-channel subcellular resolution PAM system for rapid analysis and diagnosis of tissue biopsies A. Develop a multi-channel subcellular resolution PAM system B. Develop a customized tissue biopsy holder for rapid tissue biopsy handling and imaging by PAM Specific Aim 2: Optimize and validate the multi-channel subcellular resolution PAM for translation into a robust, accurate, and efficient intraoperative device A. Optimize image acquisition in order to maximize discrimination of nuclear, cytoplasmic, and stromal properties with minimal tissue degradation B. Optimize post-acquisition algorithms to generate pseudocolored images resembling classical hematoxylin and eosin (H&E) sections with superimposed unique imaging modalities (edema and collagen) C. Validate the utilization of PAM generated images for diagnostic purposes by comparing with H&E sections, which are prepared by formalin-fixed paraffin-embedding (the current gold standard) D. Validate the utilization of PAM generated images for intraoperative diagnostic purposes by comparing with H&E sections, which are prepared by frozen section (the current intraoperative gold standard) Impact: The proposed technology will represent a major breakthrough by addressing all the limitations of current frozen section-based diagnoses and providing images with H&E equivalent qualities intraoperatively. It aims to introduce a rapid and tissue saving diagnostic method for surgical biopsies, positively impacting the healthcare system and patient care. Reducing the turnaround time on the frozen section interpretation will significantly reduce the time spent in the operating room, and therefore, decrease patient risks and costs. The ability to evaluate adequacy of small specimens with no loss in tissue will decrease the need for re-biopsy procedures. The virtually generated images can be interpreted remotely so that smaller hospitals can benefit from procedures currently available only to larger tertiary hospitals and university centers.