Chemiluminescent high-throughput microassay for evaluation of acetylcholinesterase inhibitors

Current drug therapies for Alzheimer’s disease are mainly based on acetylcholinesterase (AChE) inhibitors. However, such inhibitors may possess non-optimal pharmacological properties or cause adverse effects, therefore research for the development of new drugs is still in progress. In this paper a rapid and simple chemiluminescent assay for the in vitro evaluation of AChE inhibitors, in which the activity of AChE is measured through a series of coupled enzymatic reactions leading to light emission, is described. The assay is performed in microtiter plates, both in the 96- and 384-well formats. In the latter format up to 30 compounds can be simultaneously analyzed in a very short time (the chemiluminescence measurement lasts 5 minutes) and using small volumes of samples and reagents (the assay volume is less than 60 microL), thus allowing high throughput screening. The assay proved to be reliable, as demonstrated by the analysis of known AChE inhibitors, and the reproducibility of the results was satisfying for a screening assay (CV% in the order of 20%). It could therefore represent a valuable alternative to other methods for the preliminary evaluation of new potential AChE inhibitors.