Label-free microscopy of mitotic chromosomes using the polarization Orthogonality Breaking technique

Abstract

The vast majority of the microscopy methods currently available to study biological samples require staining prior to imaging. Nevertheless, the ability to reveal specific cell structures or organelles in a label-free manner remains desirable in different contexts. Polarization microscopy has long been considered as an interesting alternative to fluorescence-based methods in order to gain specificity on the imaged biological samples. In this work, we show how an original polarization imaging technique, implementing micro-wave photonics and referred to as orthogonality-breaking (OB) microscopy, can provide informative polarization images from a single scan of the cell sample in a fast and sensitive way. For OB imaging, the sample is probed with a laser setup simultaneously generating two orthogonal polarizations shifted in frequency by a few tens of MHz. If the imaged samples display some polarimetric properties, the orthogonality between the two polarizations is broken, leading to a beatnote interference signal that can be detected with a fast detector. The comparison of the images of various cell lines at different cell-cycle stages obtained by OB polarization microscopy and fluorescence confocal images shows that an endogenous polarimetric contrast arizes on compacted chromosomes during cell division. This technique paves the way to label-free real-time polarization imaging of mitotic chromosomes with further potential applications in histology and cancer diagnosis.