Study of Chromatin Organization by Circular Intensity Differential Scattering (CIDS)

Chromatin is the ensemble of proteins and DNA contained inside the nucleus. The study of its structure (and in particular distinguishing the more compacted heterochromatin areas from the less compacted euchromatin ones) has been of great interest owing to its role in many important processes like cell replication and illness development. While most biophysical imaging methods involve fluorescence microscopy, the introduction of markers requires additional steps and can lead to changes in the fundamental conformation of the observed structures. In my work, I have helped create and utilized a custom Circular Intensity Differential Scattering (CIDS) microscope as a label-free polarization based technique to image chromatin conformation. The CIDS signal is the scattering part of the circular dichroism and it has been shown in the past to be sensitive to chiral structures such as the ones present in chromatin and in particular to their density, radius and pitch. By comparing CIDS imaging with fluorescence imaging of isolated cell nuclei, I have shown differences in CIDS signal that have been attributed to the different density of heterochromatin and euchromatin. Finally, the technique has been improved by the coupling of CIDS imaging with Expansion microscopy in what we have called ExCIDS. Expansion Microscopy allows to artificially increase the optical resolution of a microscope by physically expanding the features of interest. The process also includes a digestion step in which biological matter that is not chromatin is eliminated, decreasing the noise due to spurious scattering and improving signal quality in ExCIDS compared to CIDS microscopy.