Solvent‐free tissue processing using supercritical carbon dioxide

Bleuel E P, Roebers T P C, Schulting E & den Dunnen W F A 
(2012) Histopathology 
 Solvent‐free tissue processing using supercritical carbon dioxide Aims:  Xylene is most often employed in tissue processing protocols for paraffin embedding, but poses a health hazard. The aim of this study was to evaluate a solvent‐free processing protocol that uses supercritical carbon dioxide (scCO 2 ) as an intermediate. Methods and Results:  A series of tests (with bovine tissues) was run, evaluating dehydration and tissue shrinkage in our new scCO 2 ‐based protocol as compared with routine processing using a graded ethanol and xylene series. A series of tests was then run to evaluate the significance of processing parameters for the outcome. Finally, a validation series was performed with optimal conditions, testing various human tissues with several staining methods. The tissue water content after paraffination was the same with our new scCO 2 ‐based protocol and the routine xylene‐based protocol. Tissue shrinkage was similar with the two methods, at ∼15%, which is also similar to values in the literature. In the validation series, the human tissues showed good morphology with strong staining, probably because of stronger antigenicity. Conclusions:  This scCO 2 ‐based protocol has been shown to be a good solvent‐free, alternative form of tissue processing. Although not the focus of this article, the time needed for tissue processing with this new protocol is within 4 h, and there is no need to change macroscopy/sectioning protocols.