Miniaturization of an Enzyme Assay (β-Galactosidase) in the 384- and 1536-Well Plate Format

Abstract

Miniaturization is one way to realize today's demands in the drug discovery process by moving from the standard 96-well plate to higher density microplate formats. In this article we describe the adaptation of a fluorescence-based enzyme assay to the challenges of the 384- and 1536-well plate format. The liquid-handling was realized by the automated micropipettor CyBi-Well™ 96/384/1536* (CyBio AG — formerly JENOPTIK Bioinstruments Gmbh — Jena, Germany). On the basis of optimized liquid-handling parameters pipetting routines were established to perform an enzyme assay (β-galactosidase) in the microplate formats of higher density. Finally, the experimental results were compared to those obtained in the well-established 96-well format. In the enzyme assay, the bioconversion of the substrate Fluorescein-di-(β-D-galactopyranoside) (FDG), occurred as a linear function of the β-galactosidase concentration comparably in all three assay formats. We conclude that miniaturization using the higher density 384- and 1536-well plate formats is advantageous as the next evolutionary step in HTS, especially using enzyme assays. A careful individual adaptation procedure for each microplate format and assay at the basis of the optimized liquid-handling parameters is essential. CyBi-Well™ 96/384/1536 proves to be a powerful tool for a careful adaptation of the liquid-handling procedures of biological assays especially also in the 384- and 1536-well formats.