Three-dimensional measurement of RBC speed, flux, and density using optical projection functional angiography

We present optical projection functional angiography (OPFA) for three-dimensional, quantitative mapping of red blood cell (RBC) speed, flux, and density independently within vascular networks based on intensity fluctuation decorrelation (IFD) and optical projection tomography (OPT). Under low coherence polarized light illumination, the intrinsic, high-contrast intensity fluctuation signal generated by moving RBCs can be effectively captured. Within the IFD framework, we define three quantitative metrics: the zero-order and first-order moments of the power density spectrum of the intensity fluctuation signal are proportional to the RBC density and flux, respectively, while the velocity is derived by dividing flux by density. Simulations and phantom experiments confirm that these decorrelation metrics enable accurate quantification of hemodynamic parameters. Multiview functional images acquired at different projection angles are combined to reconstruct the three-dimensional full-field functional angiography. The method was subsequently applied to whole-body vascular imaging in tumor-bearing zebrafish. Our findings reveal that tumor vessels exhibit reduced functional differences among different blood vessels and can be detected at an early stage of tumor development, prior to observable structural changes in the vasculature. These results suggest potential applications in early cancer diagnosis and in the study of physiological development using zebrafish disease models.