Lateral shear and digital holographic microscopy to check dynamic behaviour of biological cell

Abstract

Quantitative Phase Microscopy (QPM) by means of Lateral Shear Interferometry (LSI) is presented as a metrological tool to investigate dynamical behaviour of living cells. QPM achieved by a Digital Holographic Microscope (DHM), in transmission geometry, allows to reconstruct object shape from the numerical reconstruction of the complex field transmitted by the object. This geometry is useful in special case of biological material because they present an amplitude contrast too small for quantitative amplitude microscopy. DHM realized with a Mach-Zehnder interferometer provides the object phase map which is simply related to the object height. This technique gives a detailed map of the internal structure of the cell. The main trouble in QPM is the removal of the aberration due to the optical setup. Usually, in DHM the aberrations are eliminated by subtraction of a reference phase map (acquired without sample cell) from the object phase map. We propose a method, based on LSI, for the aberration removal that avoids this double exposure and makes use of the single object hologram. The reconstructed wavefront and its shifted replica are subtracted to obtain a shearogram from which the phase map of the object can be completely retrieved. Both shifted replica and subtraction are processed numerically. This method allows to remove optical aberrations more efficiently and faster than other conventional techniques and provides real time monitoring of cell samples.