Histochemical staining and biochemical assay of acetylcholinesterase (AChE) activity were applied to rectal suction biopsy tissue from patients treated by Z-shaped anastomosis for Hirschsprung's disease. Postoperative AChE activity and AChE/Total ChE (% AChE) in the rectal anterior wall were significantly higher (p < 0.001) than those in the posterior wall. Biochemical AChE activity was well correlative to visual mucosal AChE activity. It is remarkable that AChE activity in the rectal anterior wall after Z-shaped anastomosis was found to be kept at high level for a long period without any relation to the time interval after the operation, the ano-rectal function, and the recto-anal reflex.