Blood flow imaging in zebrafish by laser doppler digital holography

Microvessel blood flow imaging techniques are widely used in biomedical research and clinical diagnostics where many diseases have a vascular etiology or involvement. For testing purposes, zebrafish embryo provides an ideal animal model to achieve high-resolution imaging of superficial and deeply localized vessels. Moreover, the study of the formation of a closed circulatory system in vertebrates is a topic of recent interest in biophysics. However, most of the existing techniques are invasive due to the use of a contrast agent for imaging purposes. Recent developments in Digital Holography and Laser Doppler Holography techniques can be considered to alleviate this issue. Laser Doppler holography and transmission microscopy can be coupled to analyze blood flow in fish embryos by adapting a laser Doppler holographic setup to a standard bio-microscope: the two beams of the holographic interferometer (illumination of the object and reference), whose frequency offset is controlled, were addressed to the microscope by optical fibers. Multimodal acquisition and analysis of the data is made by acting on the frequency offset of the two beams, and on the location of the Fourier space filtered zone. In this work, we show that it is possible to select the signal of moving scatterers, and to image Red Blood Cells (RBCs) and blood vessels. Individual RBCs are imaged, and movies showing the RBC motion are obtained. Microsc. Res. Tech. 81:153-161, 2018. © 2016 Wiley Periodicals, Inc.