Laboratory-based cellular-level correlative visible-light and X-ray microscopy for 3D evaluation of mouse kidney...

X-ray microscopy (XRM) is an emerging technology for nondestructive evaluation of bio-specimen. We previously reported that paraffin, which is used for normal histology, acts as a tissue contrast enhancement agent in XRM, thereby enabling cellular-level 3D visualization of unstained biological samples. In this work, a new form of correlative visible-light and X-ray microscopy (CLXM) was introduced, where a kidney biopsy sample was first observed by laboratory-based XRM with paraffin-mediated contrast enhancement, and the same sample was then observed by visible-light microscopy (LM) at the same measurement point, to compare images from these two modalities precisely. This laboratory-based cellular-level CLXM was applied to a kidney biopsy from a 5/6 nephrectomy model mouse to evaluate glomerular hypercellularity lesions. The image comparison identified high-opacity particles in XRM images as cell nuclei in various parts of the nephron including glomeruli and tubules. A complementary approach using both LM and XRM images allowed us to identify/segment three lesions as aggregations of mesangial cells. It was found that one mesangial cell occupied a volume of about 100 μm³ in these lesions showing different 3D morphology. Present results suggest the availability of laboratory-based XRM for the practical 3D evaluation of whole kidney biopsies at cellular-level spatial resolution.