Quote from "The SAGES Manual of Pediatric Minimally Invasive Surgery" by Danielle S. Walsh, Todd A. Ponsky, Nicholas E. Bruns -

"Suction rectal biopsy is the gold standard for making the diagnosis. There is a normal paucity of ganglia in the region of the internal sphincter; therefore the biopsy should be taken at least 1–2 cm above the dentate line. Acetylcholinesterase staining will demonstrate hypertrophied nerve trunks and a lack of normal ganglia in classical HD. Loss of calretinin immunostaining is also consistent with the diagnosis of HD [23]."