Common-path digital holographic microscopy for 3D nanoparticle-localization

Nanoparticle localization is an important tool for a wide range of applications from biomedical imaging to fluid mechanics and particle dynamics. We present a method for three-dimensional localization of micro-and nanoparticles based on a common-path digital holographic microscope. In addition to amplitude images of conventional light microscopy, digital holography utilizes additional phase information, which allows three-dimensional representation of objects. While the lateral resolution is diffraction limited, quantitative phase measurement in combination with a novel depth-filtering technique enable an axial localization accuracy which exceeds the lateral resolution by far. This contributes to a more exact localization of particles and allows detailed characterization of structures. Our common-path interferometric setup offers high stability, which is a critical aspect in interferometry, as both reference beam and object beam follow the same optical path. Since it takes advantage of self-referencing it is also very insensitive for instabilities in the sample or sample path. The samples contain nanoparticles of varying size located in a transparent carrier material. They are placed in reflection geometry and illuminated by a diode laser at 760 nm. Their reflection is captured by a microscope objective, which provides the necessary magnification. A phase sensitive sCMOS-camera captures the image, which is then reconstructed using the angular spectrum method and a number of numerical correction methods.