This paper describes properties of microplates (MTPs) with integrated, fluorescence-based sensors for pH and oxygen, and their application for enzyme screening and monitoring of bacterial respiratory activity. Thin, hydrophilic sensing films consisting of an analyte-sensitive indicator and a reference fluorophore, are deposited on the bottom of the MTP This allows for calibration-free quantification of pH and pO(2) with an acceptable accuracy and resolution for applications such as enzyme activity screening, respirometry, or toxicological assays. The sensor properties are mainly investigated with respect to their qualification for such applications. Specifically, enzyme activity screening is demonstrated for glucose oxidase using oxygen-sensitive plates, and for bacterial growth monitoring of Escherichia coli and Pseudomonas putida. Furthermore, a toxicological assay monitoring the respiration activity of P. putida was converted into a microplate format. (c) 2005 Elsevier B.V. All rights reserved.