Large depth-of-field tracking of colloidal spheres in holographic microscopy by modeling the objective lens

Abstract

Holographic microscopy has developed into a powerful tool for 3D particle tracking, yielding nanometer-scale precision at high frame rates. However, current particle tracking algorithms ignore the effect of the microscope objective on the formation of the recorded hologram. As a result, particle tracking in holographic microscopy is currently limited to particles well above the microscope focus. Here, we show that modeling the effect of an aberration-free lens allows tracking of particles above, near, and below the focal plane in holographic microscopy, doubling the depth of field. Finally, we use our model to determine the conditions under which ignoring the effect of the lens is justified, and in what conditions it leads to systematic errors.