Ratzlaff E H, Grinvald A
IBM Research Division, Yorktown Heights, NY 10598.
J Neurosci Methods. 1991 Feb;36(2-3):127-37. doi: 10.1016/0165-0270(91)90038-2.
The design of a macroscope constructed with photography lenses is described and several applications are demonstrated. The macroscope incorporates epi-illumination, a 0.4 numerical aperture, and a 40 mm working distance for imaging wide fields in the range of 1.5-20 mm in diameter. At magnifications of 1X to 2.5X, fluorescence images acquired with the macroscope were 100-700 times brighter than those obtained with commercial microscope objectives at similar magnifications. In several biological applications, the improved light collection efficiency (20-fold, typical) not only minimized bleaching effects, but, in concert with improved illumination throughput (15-fold, typical), significantly enhanced object visibility as well. Reduced phototoxicity and increased signal-to-noise ratios were observed in the in vivo real-time optical imaging of cortical activity using voltage-sensitive dyes. Furthermore, the macroscope has a depth of field which is 5-10 times thinner than that of a conventional low-power microscope. This shallow depth of field has facilitated the imaging of cortical architecture based on activity-dependent intrinsic cortical signals in the living primate brain. In these reflection measurements large artifacts from the surface blood vessels, which were observed with conventional lenses, were eliminated with the macroscope.
本文描述了一种用摄影镜头构建的宏观显微镜的设计，并展示了其几种应用。该宏观显微镜采用落射照明，数值孔径为0.4，工作距离为40 mm，用于对直径在1.5 - 20 mm范围内的宽视野进行成像。在1倍至2.5倍的放大倍数下，用该宏观显微镜采集的荧光图像比在类似放大倍数下用商业显微镜物镜获得的图像亮100 - 700倍。在几个生物学应用中，提高的光收集效率（通常为20倍）不仅将漂白效应降至最低，而且与提高的照明通量（通常为15倍）协同作用，还显著增强了物体的可见性。在使用电压敏感染料对皮质活动进行体内实时光学成像时，观察到光毒性降低且信噪比提高。此外，该宏观显微镜的景深比传统低倍显微镜薄5 - 10倍。这种浅景深有助于基于活体灵长类大脑中活动依赖的内在皮质信号对皮质结构进行成像。在这些反射测量中，宏观显微镜消除了用传统镜头观察时出现的来自表面血管的大量伪影。
