Background
Confocal Scanning Laser Microscopy (CSLM) is an advanced, non?invasive imaging technique originally developed for in vivo applications and increasingly used ex vivo without compromising cytological or tissue integrity. Reflectance Confocal Microscopy (RCM) provides high?resolution visualization of cellular structures by exploiting differences in refractive indices, enabling real?time “virtual biopsy” imaging comparable to hematoxylin–eosin sections. Within pathology workflows, CSLM offers rapid assessment of sample adequacy while preserving the entire specimen for downstream immunohistochemical and molecular analyses.
Objectives
This study assessed the applicability of laser confocal microscopy (VIVASCOPE^TM 2500) for rapid ex vivo evaluation of serous effusions, with objectives including real?time adequacy assessment, preliminary morphological interpretation, full material preservation for ancillary testing, and implementation of a standardized digital alternative to traditional cell?block.
Methods
Thirty abdominal ascitic and pleural effusion samples were received as fresh specimens and processed in duplicate to enable direct methodological comparison. For each case, one aliquot was prepared on a Cytomatrix support and scanned with the VIVASCOPE^TM 2500 to generate a pseudo?H&E for confocal evaluation, while the paired aliquot underwent standard cytological inclusion to obtain a conventional effusion H&E from cell?block pellet. This dual?processing strategy allowed comparison between digital pseudo? H&E and Cytomatrix?derived H&E to assess sample architecture and material recovery, and between Cytomatrix?based H&E and H&E from routine cyto?inclusion to evaluate morphological preservation. All Cytomatrix?embedded samples were fully recovered and processed for histological, immunohistochemical, and molecular analyses to confirm material retention and downstream compatibility.
Results
Confocal microscopy generated high?resolution images up to ~100 µm depth, enabling clear visualization of cellular and subcellular structures. Digital slides were comparable to H&E sections and allowed rapid assessment of sample adequacy in all cases. Cytomatrix preparation ensured homogeneous cell distribution and prevented material loss, in contrast to cell?block processing, where low?cellularity samples often yield suboptimal pellets. Morphological preservation on Cytomatrix? VIVASCOPE^TM 2500 slides was superior, with intact nuclear detail, crisp cytoplasmic borders, and well?maintained architectural patterns, whereas cell?block sections occasionally showed artefactual distortion or fragmentation. All Cytomatrix?embedded specimens were successfully processed for immunohistochemical and molecular analyses, confirming full material preservation and compatibility with ancillary testing.
Conclusions
Laser confocal microscopy using the VIVASCOPE^TM 2500, combined with Cytomatrix preparation, offers a robust digital modality for rapid cytological evaluation of serous effusions. It enables real?time “virtual biopsy” visualization, ensures complete material preservation without the quantitative loss typical of cell?block processing, and maintains superior morphological integrity. This workflow provides a safe, reproducible alternative to cell?block preparation and enhances diagnostic efficiency while supporting the broader adoption of digital pathology technologies.