Whole-mount NADPH-diaphorase histochemistry is a reliable technique for the intraoperative evaluation of extent of aganglionosis

Multiple seromuscular biopsies at three levels (narrow segment, transitiona l zone, and dilated segment) were taken and investigated intraoperatively t o determine the extent of aganglionosis. Using the wholemount preparation t echnique, circular muscle fibers were separated from the specimens. After a short prefixation, the muscle fibers were stained by the NADPH-diaphorase technique and were examined within 20-25 min. A fine and dense neuronal mes hwork was observed between circular muscle fibers in the normal and ganglio nic part of the bowel. In contrast, there was a complete lack of NADPH-diap horase-positive fibers in the circular muscle of aganglionic colon. In the transitional zone, NADPH-diaphorase-positive fibers were markedly reduced c ompared to the ganglionic region. The density of these fibers increased and attained normal levels in the proximal bowel above the transition zone. Th ese results suggest that whole-mount NADPH-diaphorase histochemistry is a t hree-dimensional technique suitable for the intraoperative evaluation of ex tend of aganglionosis. The technique is sufficiently rapid to be used in co njunction with routine frozen sections to assist in the diagnosis and in se lecting the optimal level of resection at the time of pull-through operatio n.