Visualizing fixed cells and tissues only gives snapshots of cellular processes. To have a better insight into dynamic events ocurring in the cells or to vizualize interactions between various cellular components in real time (e.g., proteins, organelles, second messengers…), powerful microscopic approaches have been developed over the past decade. This post will review the recent live cell imaging probes developed by Goryo Chemicalsand available in Europe through tebu-bio.
Cell-based calcium assays have been a mainstay of GPCR (G protein coupled receptor) drug discovery and signal transduction research since the mid 1990s. However, a calcium readout alone might be ambiguous, because many cellular processes produce calcium. Furthermore, it does not differentiate between signalling pathways routed through the different types of heterotrimeric G proteins. While effects on GPCRs which exhibit effects on Gs proteins lead to an elevated level of the second messaeger cAMP by activating the activity of adenylate cyclase, signalling through Gi proteins inhibits adenylate cyclase and thus leads to a decrease of cAMP. Finally signalling through Gq proteins leads to phospolipase C activation which cleaves Phosphatidylinositol 4,5-bisphosphate (PIP2) to Diacetlyglycerol (DAG) and Inositol trisphosphate (IP3) which leads to an increase in Ca2+ level (for an overview see reference 1).
Dye based methods to measure Ca2+ are very well established, whereas there are no dyes available for most of the other second messengers in these signalling pathways.
Genetically encoded fluorescent biosensors for live cell discovery
That’s why Montana Molecular developed genetically encoded fluorescent biosensors for second messengers such as cAMP, DAG, PIP2, and Ca2+, references 2, 3). [Read more…]
G Protein Coupled Receptors (GPCR) and ion channels are important targets in drug development. Both target classes represent membrane proteins which require convenient and robust cell-based assays for screening campaigns. Usually used as readout of such screenings, intracellular Calcium concentration implies the use of Ca2+ indicators able to meet demanding expectations of researchers engaged in assay development and screenings. [Read more…]