Lee Grace S, Miele Lino F, Turhan Aslihan, Lin Miao, Hanidziar Dusan, Konerding Moritz A, Mentzer Steven J
Laboratory of Adaptive and Regenerative Biology, Brigham and Women's Hospital, Department of Surgery, Harvard Medical School, Boston, Massachusetts 02115, USA.
Microsc Res Tech. 2009 Feb;72(2):85-92. doi: 10.1002/jemt.20647.
Quantitative assessment of microvascular structure is relevant to the investigations of ischemic injury, reparative angiogenesis and tumor revascularization. In light microscopy applications, thick tissue specimens are necessary to characterize microvascular networks; however, thick tissue leads to image distortions due to out-of-focus light. Structured illumination confocal microscopy is an optical sectioning technique that improves contrast and resolution by using a grid pattern to identify the plane-of-focus within the specimen. Because structured illumination can be applied to wide-field (nonscanning) microscopes, the microcirculation can be studied by sequential intravital and confocal microscopy. To assess the application of structured illumination confocal microscopy to microvessel imaging, we studied cell-sized microspheres and fused silica microcapillary tissue phantoms. As expected, structured illumination produced highly accurate images in the lateral (X-Y) plane, but demonstrated a loss of resolution in the Z-Y plane. Because the magnitude of Z-axis distortion was variable in complex tissues, the silica microcapillaries were used as spatial calibration standards. Morphometric parameters, such as shape factor, were used to empirically optimize Z-axis software compression. We conclude that the silica microcapillaries provide a useful tissue phantom for in vitro studies as well as spatial calibration standard for in vivo morphometry of the microcirculation.
微血管结构的定量评估与缺血性损伤、修复性血管生成和肿瘤血管再生的研究相关。在光学显微镜应用中，需要厚组织标本才能表征微血管网络；然而，厚组织会因离焦光而导致图像失真。结构照明显微镜是一种光学切片技术，它通过使用网格图案来识别标本内的焦平面，从而提高对比度和分辨率。由于结构照明可应用于宽视场（非扫描）显微镜，因此可以通过连续活体显微镜和共聚焦显微镜来研究微循环。为了评估结构照明显微镜在微血管成像中的应用，我们研究了细胞大小的微球和熔融石英微毛细管组织模型。正如预期的那样，结构照明在横向（X-Y）平面上产生了高度精确的图像，但在Z-Y平面上显示出分辨率的损失。由于在复杂组织中Z轴失真的程度是可变的，因此使用二氧化硅微毛细管作为空间校准标准。形态测量参数，如形状因子，被用于凭经验优化Z轴软件压缩。我们得出结论，二氧化硅微毛细管为体外研究提供了有用的组织模型，也为微循环的体内形态测量提供了空间校准标准。
