Pomelo (Citrus maxima) as a Clearing Extract in Tissue Processing

Abstract

Clearing is a critical step in tissue processing which involves removing alcohol from tissue samples to prepare them for paraffin embedding, in a routine tissue processing. Xylene is volatile, readily producing flammable and toxic concentrations at room temperature. A known non-toxic substance with similar characteristics with xylene is called D-limonene. It appears to perform adequately as a wax solvent and cleaning agent, as it has a reduced fire risk compared with xylene. In this study, D-Limonene from pomelo peel extract was utilized as a clearing agent in tissue processing. This study aims to: 1) determine the effect of pomelo peel extract on chicken liver tissue; 2) investigate the most effective clearing time of pomelo peel extract; and 3) compare the effectiveness of different treatments against xylene. The control group consisted of 3 replicates of chicken liver tissue cleared with xylene, while the experimental group included 3 treatments; treatment 1 (60 minutes), treatment 2 (120 minutes), and treatment 3 (180 minutes) with 3 replicates each. Statistical analysis revealed that the mean of treatment 2 indicated a good clearing agent in terms of clearing ability through evaluation of the translucency, evaporation rate in the paraffin oven, compatibility with the hematoxylin and eosin stain, morphological changes of the chicken liver tissue based on the degree of shrinkage or swelling, while the control group showed a satisfactory clearing ability. These results indicate the potential of pomelo (Citrus maxima) peel extract as an alternative clearing agent in tissue processing. Therefore, further studies are strongly recommended.