Wieringa P A, Damon D N, Duling B R
Department of Physiology, University of Virginia School of Medicine, Charlottesville 22908, USA.
Med Biol Eng Comput. 1995 Jul;33(4):563-70. doi: 10.1007/BF02522515.
A method has been developed to examine the movement of plasma in capillaries using intravital microscopy. Spatial transients in fluorescence properties are instantaneously induced by laser photo-bleach pulses after which the convective recovery can be monitored. The plasma is tagged with fluorescent dyes coupled to bovine serum albumin, which is injected well before the measurements and circulates with the blood stream. A laser beam from an argon laser source, set to emit light with a wavelength of 488 nm, is focused on the illumination field diaphragm and creates a spot in the object plane of the microscope. At low laser power, the laser spot is aimed at a blood plasma gap between red blood cells in a capillary segment, using a steerable mirror. Light sensors, coupled to photo-multipliers in the secondary image plane of the microscope, record the light intensity of the moving plasma/dye while the preparation is continuously illuminated with a xenon epi-illuminating set-up. The laser photo-bleach spot is then used to bleach the dye complex within a 5.4 microns segment of the capillary for less than 20 ms. The movement of the bleached plasma bolus is tracked by the photo-sensors, placed sequentially along the capillary. Both dye and red blood cell passage can be detected in the photo-multiplier signals, and the relative velocities of the two blood components can be measured. Measurements reveal that the ratio of transit times between blood plasma and red blood cells is 1.23 (SD = 0.22, N = 18), which is in good agreement with measurements by other techniques.
已开发出一种利用活体显微镜检查毛细血管中血浆流动的方法。荧光特性的空间瞬变由激光光漂白脉冲瞬间诱发，之后可监测对流恢复情况。血浆用与牛血清白蛋白偶联的荧光染料标记，在测量前很久就进行注射，并随血流循环。来自氩激光源的激光束设置为发射波长为488 nm的光，聚焦在照明场光阑上，并在显微镜的物平面上形成一个光斑。在低激光功率下，使用可操纵镜子将激光光斑对准毛细血管段中红细胞之间的血浆间隙。与显微镜二次图像平面中的光电倍增管相连的光传感器，在使用氙灯落射照明装置持续照明标本的同时，记录移动的血浆/染料的光强度。然后用激光光漂白光斑在不到20毫秒的时间内漂白毛细血管5.4微米段内的染料复合物。沿毛细血管依次放置的光传感器跟踪漂白血浆团块的移动。在光电倍增管信号中可检测到染料和红细胞的通过情况，并可测量两种血液成分的相对速度。测量结果显示，血浆和红细胞的通过时间之比为1.23（标准差=0.22，N=18），这与其他技术的测量结果高度一致。
