Related Experiment Video
Updated: Jul 20, 2026
A Fluorescence-based Lymphocyte Assay Suitable for High-throughput Screening of Small Molecules
Published on: March 10, 2017
High-throughput screening for monoamine oxidase-A and monoamine oxidase-B inhibitors using one-step fluorescence
1Institute of Materia Medica, Peking Union Medical College and Chinese Academy of Medical Sciences, Beijing 100050, China.
Aim:
To develop high-throughput screening (HTS) assays for monoamine oxidase (MAO)-A and MAO-B inhibitors.
Methods:
A fluorescence probe based method measuring MAO-A and MAO-B activity was established and optimized, with its sensitivity, stability and specificity evaluated. Reaction conditions including enzyme sources, substrate concentrations, incubation volume and reaction time in 384-well format were optimized to achieve sensitive and low consumptive goal.
Results:
In optimized conditions, dynamic parameters of MAO-A and MAO-B were obtained. The K(m) value of serotonin to MAO-A was 1.66 micromol/L, while that of benzylamine to MAO-B was 0.80 micromol/L. The IC(50) value of clorgyline to MAO-A was 2.99 nmol/L, and that of deprenyl to MAO-B was 7.04 nmol/L, matching those obtained from traditional spectrometric assays. Among tested samples, one compound exerted an inhibitory effect on MAO-A activity with IC(50) as 0.36 micromol/L, and three compounds had an inhibitory effect on MAO-B activity with IC(50) as 0.13, 0.19, and 0.13 micromol/L. The Z' factor was 0.71+/-0.03 and 0.75+/-0.03 in MAO-A-inhibitor and MAO-B-inhibitor HTS system, respectively.
Conclusion:
The established assays can be well applied to MAO-A and MAO-B inhibitor screening with high quality, precision and reproducibility.
