Enzymatic microdetermination method for choline acetyltransferase

An enzymatic microdetermination method for choline acetyltransferase was developed by measuring acetyl-CoA formed in the reverse reaction and applied to analysis of single neurons. Cell bodies of single neurons were isolated from freeze-dried sections of rabbit spinal cords and weighed on a highly sensitive quartz fiber balance. These cell body samples (0.6-3 ng) were added directly to the reaction mixture (13~-54 nl). The acetyl-CoA formed was amplified 64,000 to 420,000 times by enzymatic amplification reactions, i.e., CoA and NADP cyclings, and determined fiuorometrically. The average activity of single anterior horn cells was 45-fold higher than that of rabbit cerebral cortex and the specific activity on a dry weight basis was widely distributed among individual neurons. No activity was detected in the non-cholinergic dorsal root ganglion cells.