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Expansion microscopy (ExM) is a novel preparation method enhancing the optical resolution by expanding uniformly the relative distance between fluorescence molecules on a sample placed inside a polymerized gel matrix. However, a skilled operator is needed for fluorescent labeling protocols and a high light dose is required for measurement. In this work, we couple ExM with a label-free differential circular polarization microscopy technique, demonstrated to be sensitive to the chiral organization of biopolymers. We show that by improving the distance between chiral groups, the new imaging contrast gives access to a better resolution of the chromatin-DNA organization in situ.

Circular Intensity Differential Scattering (CIDS) provides a differential measurement of the circular right and left polarized light and has been proven to be a gold standard label-free technique to study the molecular conformation of complex biopolymers, such as chromatin. In early works, it has been shown that the scattering component of the CIDS signal gives information from the long-range chiral organization on a scale down to 1/10th–1/20th of the excitation wavelength, leading to information related to the structure and orientation of biopolymers in situ at the nanoscale. In this paper, we review the typical methods and technologies employed for measuring this signal coming from complex macro-molecules ordering. Additionally, we include a general description of the experimental architectures employed for spectroscopic CIDS measurements, angular or spectral, and of the most recent advances in the field of optical imaging microscopy, allowing a visualization of the chromatin organization in situ.