An X-ray micro-CT protocol enables non-destructive 3D visualization of glandular architecture in archived paraffin-embedded prostate cancer tissue.

The prostate is organized as a complex glandular network, and prostate cancer is accompanied by architectural changes that cannot always be inferred from two-dimensional (2D) histology. The study of these features in three dimensions (3D) in cancerous tissue is important because glandular features have shown potential value for disease characterization and risk stratification beyond that obtained from individual 2D sections. Large collections of clinically annotated material preserved in paraffin remain an important research source. However, many 3D imaging methods require destructive or labor-intensive preparation, limiting access to valuable material that needs to be re-archived. In this proof-of-concept study, we developed an X-ray micro-computed tomography (micro-CT) workflow for direct 3D imaging of archived paraffin-embedded prostate specimens with near-zero sample manipulation. Imaging and denoising parameters were optimized to balance image quality with acquisition time. Paraffin-embedded prostate core punches (n = 5) from five radical prostatectomy specimens were imaged by micro-CT and subsequently processed for histology. Digitized histological sections were aligned with corresponding micro-CT planes for qualitative comparison, while pre-sectioning micro-CT data provided an intact geometric reference for exploratory assessment of histology-associated dimensional changes at the whole-core and glandular-lumen levels. The optimized workflow yielded micro-CT images with discernible glandular morphology and qualitative structural correspondence with matched histological sections. Exploratory measurements suggested modest dimensional change at the whole-core level and greater variability at the lumen level; none of the tested deformation metrics reached conventional statistical significance. Illustrative 3D segmentation presented the glandular epithelium and lumina as spatially continuous structures within a cancer-containing region. These findings support the feasibility of X-ray micro-CT as a non-destructive approach for 3D characterization of prostate glandular architecture directly from paraffin-embedded tissue, providing volumetric context for structures identified on histology while preserving the specimen for subsequent histological assessment.