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Intravital molecular tagging velocimetry of cerebral blood flow using Evans Blue

The effects of light‐driven enhancement of Evans Blue dye complexes with blood plasma proteins were observed for the first time, both in vitro and in vivo. The possible background of the effect concerns the photochemical cis‐trans isomerization of the azo dye molecules. The effect was induced in the solution with a red laser with a wavelength of 638 nm, which corresponds to the peak of the dye absorption. The lifetime of the enhanced fluorescence is approximately 1 second and enables its use as an optically tagged molecular flow tracer for blood flow velocity measurements. Utilizing the effect, we performed for the first time the intravital molecular tagging velocimetry of the blood velocity in blood vessels in a living animal. The results of the measurements of the blood flow velocities in the cerebral veins of a group of healthy mice are presented.