Methods to improve the preservation of peritoneal tissues

Peritoneal mesothelial cells are easily detached during conventional tissue processing, which may result in artifacts in peritoneal tissue examination. Therefore, in the present study, we investigated several methods to improve the preservation of the anterior parietal peritoneal tissues. Peritoneal tissue from the anterior abdominal wall was taken from each of 5 rats killed for the experiment. Tissue samples were immediately treated by one of these methods: (1) fixed with 10% formaldehyde; (2) fixed with Bouin's solution; (3) fixed with Helly's solution. After fixation, the samples were dehydrated with one of (a) ethanol, 1 hour in each step; (b) ethanol, 15 minutes in each step; or (c) tertiarybutyl alcohol. Five sections were taken from each tissue and stained with hematoxylin and eosin. The quality of tissue fixation was evaluated by image analysis. Peritoneal mesothelial cells were well preserved after fixation with Helly's solution or Bouin's solution. With 10% formaldehyde, about 40% of the mesothelial cells were lost. Dehydration with ethanol--especially long-duration dehydration--increased the loss. However, dehydration with tertiarybutyl alcohol avoided the increased loss of mesothelial cells. The submesothelial extracellular matrix was well preserved with Bouin's solution, but not with the other fixatives. Our results suggest that fixation with formaldehyde and dehydration with ethanol results in significant loss of peritoneal mesothelial cells and submesothelial extracellular matrix in peritoneal tissues. Fixation with Bouin's solution and dehydration with tertiary-butyl alcohol may be a better method of preserving peritoneal tissue.