Development of Instrumentation for Fluorescence-Guided Surgery

荧光引导手术器械的开发

• 批准号: 7967908
• 负责人: Paul Smith
• 金额: $ 4.52万
• 依托单位: NATIONAL INSTITUTE OF BIOMEDICAL IMAGING AND BIOENGINEERING
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7967908
• 关键词: Animal Model; Animals; Biological Preservation; Development; Diagnostic; Dose; Drops; Excision; Feedback; Fluorescence; Fluorescent Probes; Future; Image; Imagery; Label; Lead; Light; Malignant neoplasm of cervix uteri; Malignant neoplasm of ovary; Medical; Modeling; Motion; Mus; Neoplasm Metastasis; Operative Surgical Procedures; Optics; Peritoneal; Property; Real-Time Systems; Resolution; Rhodamine; Rhodamines; Signal Transduction; Specificity; Speed; System; Techniques; Time; Tissues; Work; charge coupled device camera; clinical application; cost; data acquisition; design; fluorescence imaging; fluorophore; improved; in vivo; instrument; instrumentation; interest; nanocrystal; nanoparticle; novel; prototype; research study; success; technique development

The rapid development of molecularly targeted probes for use in vivo has led to growing interest in clinical applications that incorporate information from these probes. In particular, there is a need for techniques to visualize these targeted probes during surgery, particularly to identify tissue either for removal or preservation. When looking at fluorescent probes in tissue, the major difficulty is usually separating the probe fluorescence from the tissue autofluorescence. Because the autofluorescence has different spectral properties than the fluorophore, it can easily be separated using multispectral imaging, in which several full emission spectra can be taken for each pixel. Although the implementation of acousto-optic tunable filters has greatly increased the speed of these techniques, the minimum time required to acquire an image cube is still several seconds with commercially available systems. When imaging using fluorescent probes with lower efficiency or when used at lower doses, the image acquisition time can be as long as several minutes. This data acquisition rate makes the use of multispectral imaging to provide real-time feedback during a surgical procedure impractical. Even when used for diagnostic purposes, the unavoidable motion of the subject during the time required to acquire an image cube can lead to an unacceptable loss of spatial resolution. This project focuses on the development of instrumentation for incorporating fluorescent and multi-spectral imaging into surgical applications, using two major approaches. The first approach is to develop a system for real-time visualization of fluorescent probes to guide surgery. The second approach is to develop a system for diagnostic applications, to provide a white-light stack of images for spatial registration of the multispectral image cube. As an alternative to multispectral imaging, aggressive filtering of both emission and excitation light can help to minimize the effects of tissue autofluorescence. Although the signal level drops as the spectral window is narrowed, this can be overcome by using a sensitive camera, such as a cooled CCD, ICCD, or even an EMCCD. Because a single spectral window is used, substantially faster data acquisition is possible. The immediate application for this instrument was the identification of peritoneal metastases in ovarian cancer, using a GSA-Rhodamine Green probe developed in NCI. The first prototype instrument, used for mock cytoreductive surgery in a murine model of ovarian cancer, had sufficient sensitivity to identify labeled metastases at an image acquisition rate of five frames per second with specificity comparable to that achieved with a multispectral system. An improved prototype, which permitted image acquisition at fifteen frames per second with enhanced sensitivity, was used for cytoreductive surgery on anesthetized animals with similar success to the previous mock surgery. This year, we also continued development on a separate multispectral system to provide a white light image stack for physical registration of the multispectral image cube, using a 92:8 beam splitter and a low-cost monochrome CCD camera in parallel with the multispectral instrumenation. Preliminary experiments on a cervical cancer animal model are underway. The hardware developed for this application could be easily adapted to incorporate a second camera into the instrument for fluorescence guided surgery; this second camera could be used either for a second fluorescence image, in order to provide a simple autofluorescence correction, or for a pseudo-white light image of the surgical field. Finally, some initial proof-of-principle in vivo imaging experiments were performed using upconverters, in this case upconverting nanocrystals that absorb 980 nm light and emit in the visible to near-infrared. These novel compounds could allow single-wavelength imaging with no detectable autofluorescence; the tissue background is so low that a white light image will likely be required to provide anatomical reference. Although considerable work remains in developing these nanoparticles for medical applications, this is a promising avenue for future efforts.

用于体内的分子靶向探针的快速发展导致了对结合这些探针的信息的临床应用的日益增长的兴趣。具体地说，需要在手术期间可视化这些靶向探针的技术，特别是识别需要切除或保存的组织。当观察组织中的荧光探针时，主要的困难通常是将探针荧光与组织自身荧光分开。由于自发荧光具有与荧光团不同的光谱性质，因此可以很容易地使用多光谱成像来分离它，在多光谱成像中，每个像素可以获得几个完整的发射光谱。虽然声光可调滤光器的实现大大提高了这些技术的速度，但在商业上可用的系统中，获取图像立方体所需的最短时间仍然是几秒钟。当使用效率较低的荧光探针进行成像或使用较低剂量时，图像采集时间可能长达几分钟。这种数据采集率使得在外科手术过程中使用多光谱成像来提供实时反馈是不现实的。即使在用于诊断目的时，对象在获取图像立方体所需的时间内不可避免的运动也可能导致不可接受的空间分辨率损失。 这个项目的重点是开发用于将荧光和多光谱成像结合到外科应用中的仪器，使用两种主要方法。第一种方法是开发一个实时可视化的荧光探针系统来指导手术。第二种方法是开发一个用于诊断应用的系统，为多光谱图像立方体的空间配准提供白光图像堆叠。 作为多光谱成像的替代方法，积极过滤发射光和激发光可以帮助将组织自发荧光的影响降至最低。虽然信号电平随着光谱窗口变窄而下降，但这可以通过使用敏感的相机来克服，例如冷却的CCD、ICCD，甚至EMCCD。由于使用了单个光谱窗口，因此可以获得更快的数据。这种仪器的直接应用是使用NCI开发的GSA-罗丹明绿探针识别卵巢癌的腹膜转移。第一台原型仪器用于模拟卵巢癌小鼠模型的细胞减少手术，在每秒5帧的图像采集速率下具有足够的灵敏度来识别标记的转移瘤，其特异性可与多光谱系统相媲美。一种改进的原型，允许以每秒15帧的速度获取图像，并提高了灵敏度，用于麻醉动物的细胞减少术，取得了与之前的模拟手术类似的成功。 今年，我们还继续开发单独的多光谱系统，为多光谱图像立方体的物理配准提供白光图像堆栈，使用92：8分束器和与多光谱仪器并行的低成本单色CCD相机。宫颈癌动物模型的初步实验正在进行中。为这一应用开发的硬件可以很容易地改装成将第二个相机结合到用于荧光引导手术的仪器中；该第二个相机可以用于第二个荧光图像，以便提供简单的自体荧光校正，或者用于外科领域的伪白光图像。 最后，使用上变频器进行了一些初步的活体成像实验，在这种情况下，上变频器将吸收980 nm光并在可见光到近红外发射的纳米晶体上转换。这些新化合物可以进行单波长成像，没有可检测到的自发荧光；组织背景如此之低，可能需要白光图像来提供解剖学参考。尽管在开发这些用于医疗应用的纳米粒子方面仍有大量工作要做，但这是未来努力的一条很有希望的途径。