Cholinesterase histochemistry in the human brain: effect of various fixation and storage conditions

Human brains fixed in 3 conventional solutions were used to compare acetylcholinesterase and butyrylcholinesterase labeling after being processed by the same histochemical procedure. The best fixative was found to be 4% paraformaldehyde in a 0.1 M phosphate buffer for 24-42 h. The addition of 2% glutaraldehyde to 2% paraformaldehyde for the same fixation time reduced the staining intensity of both enzymes. Formalin after 2 days resulted in a lighter staining intensity of fibers, senile plaques and neurofibrillary tangles, whereas after 10 days it caused the near disappearance of acetylcholinesterase-positive fibers. Finally, after more than 10 days in formalin, a certain number of senile plaques and a few neurofibrillary tangles could be observed, and those only in the case in which the substrate concentration and incubation time were increased. The fixed tissue, in either blocks or sections, can be stored for more than 1 year in a glycol solution at -17 degrees C, without the inconvenience of freezing, with good histology and histochemical preservation of both enzymes.