Red cell velocity and microvessel diameter measurement by a two fluorescent tracer method under epifluorescence...

Fluorescein isothiocyanate (FITC)-labeled red blood cells (RBCs) and rhodamine-B isothiocyanate (RITC)-labeled dextran were used as two fluorescent tracers under intravital fluorescence microscopy. RBC velocity in cerebral microvessels of cats was measured with a dual window technique using FITC-labeled RBCs as a flow tracer. Measurement of the vessel diameter was performed using a digital image processor system by staining plasma with RITC-labeled dextran. The RBC velocity, obtained directly with digital cross-correlation of videodensitometric signals derived from the two windows, coincided with those obtained with the frame-by-frame analysis. The obtained RBC velocity ranged between 12.9 and 0.45 mm/sec in arterioles (< 60 microns) in diameter, 4.2 and 0.18 mm/sec in venules (< 60 microns in diameter), and 1.1 and 0.26 mm/sec in capillaries. The use of the labeled RBCs as the flow tracer enabled us to measure a wide range of RBC velocity (up to about 7 mm/sec) by setting the distance between the two windows beyond about 200 microns.