Quantitative three-dimensional imaging of droplet convection and cardiac cell motions based on micro DDPIV.

Abstract

Biomechanical forces such as blood flow induced shear stress as well as genetic programming are widely acknowledged as critical factors regulating vertebrate heart development. While mechanisms of genetic regulation have been well studied, effects of biomechanics are poorly understood due to the lack of proper imaging tools with sufficient spatial and temporal resolutions for quantitative analysis of the mechanical stimuli in complex three-dimensional (3D) living systems. 3D quantitative flow visualization by tracking microscale particles has become an invaluable tool in microfluid mechanics. Defocusing digital particle image velocimetry (DDPIV) can recover depth coordinates by calculating the separation between defocused images generated by an aperture mask with a plurality of pinholes forming an equilateral triangle. In this thesis, a novel high-speed 3D micro-PTV system was developed based on this technique with laser-induced fluorescence to achieve microscale velocity field measurements. Application of this technique to microscale imaging was validated by calibration of targets spread over the image field. A micro volume of 400x300 µm2 with 100 µm depth has been mapped using an inverted microscope equipped with a 20X objective lens. The proposed technique was successfully applied to 3D tracking of 2-µm fluorescent particles inside an evaporating water droplet, exhibiting convective flow induced by Marangoni effects. The microscopic imaging system was then utilized to acquire 3D time series data of highly dynamic cell motions in living embryonic zebrafish hearts. 1-µm and 500-nm fluorescent tracer particles were injected into the blood stream of developing zebrafish embryos at 32 hours post fertilization (hpf) to 59 hpf to help describe cardiac cell motions. Microinjection was delicately performed at the fish tail to minimize the influence to normal cardiovascular functions. The measurable depth in an embryonic heart is about 40 µm. 3D velocities of cardiovascular blood flow and trajectories of heart-wall motions were obtained, showing dynamic changes of the flow field and phase differences of wall movements between the atrium and the ventricle during heart beating. Endocardial ventricular strains were calculated based on the reconstructed coordinates of two particles adhered to the endocardium.