Indicator displacement assay using an <i>in situ</i> generated polymeric system in water: exploiting donor–acceptor interactions

<p>Indicator displacement assays (IDAs) using an electron-rich polymer as the receptor for electron-deficient guests are described in this study. The electron-rich water-soluble polymeric system possesses hydrophobic pockets, which were generated by the reaction of <b>1</b> (a Michael donor) and 1,3,5-triacryloylhexahydro-1,3,5-triazine, <b>5</b> (TAT, a Michael acceptor), in water at pH 8.2 in 2 min at 85^oC. When electron-deficient fluorescent indicators are added to the polymeric systems, large changes in fluorescent intensity of the indicators occur. This donor–acceptor architecture between polymer and indicator was in turn explored for displacement of the indicator with electron-deficient analytes such as TNT.</p

In-Situ Generation of Differential Sensors that Fingerprint Kinases and the Cellular Response to Their Expression

Mitogen-activated protein (MAP) kinases are responsible for many cellular functions, and their malfunction manifests itself in several human diseases. Usually, monitoring the phosphorylation states of MAP kinases in vitro requires the preparation and purification of the proteins or Western blotting. Herein, we report an array sensing approach for the differentiation of MAP kinases and their phosphorylated counterparts in vitro. This technique utilizes a library of differential receptors created in situ containing peptides known for affinity to MAP kinases, and a Zn­(II)–dipicolylamine complex that binds phosphate groups on proteins. An indicator-displacement assay signals the binding of the individual receptors to the kinases, while chemometrics is used to create a fingerprint for the kinases and their state of activity. For example, linear discriminant analysis correctly identified kinase activity with a classification accuracy of 97.5% in vitro, while the cellular response to kinase expression was classified with 100% accuracy