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Beschreibung

Millipore's Ready-to-Assay GPCR frozen cells are designed for simple, rapid calcium assays with no requirement for intensive cell culturing. Millipore has optimized the freezing conditions to provide cells with high viability and functionality post-thaw. The user simply thaws the cells and resuspends them in media, dispenses cell suspension into assay plates and, following over night recovery, assays for calcium response. Lysophosphatidic acid (LPA) is a lysophospholipid produced by activated platelets that inhibits adenylate cyclase and stimulates DNA synthesis, changes in cell morphology, and increases in intracellular calcium in a variety of cultured mammalian cells. A growing family of GPCRs mediates the biological effects of LPA. The first LPA receptors described, LPA1-3, share relatively high sequence similarity and are related to the sphingosine 1-phosphate receptors S1P1-5 (Contos et al., 2000). Two more recently characterized LPA receptors, LPA4 and LPA5, are related to each other but are more distantly related to LPA1-3. LPA5, originally known as GPR92, mediates LPA-induced cytoskeletal changes, intracellular calcium flux and increased cAMP by coupling to G12/13, Gq, and Gs. (Lee et al., 2006). In addition to binding LPA, LPA5 is also activated by several other lipid-derived molecules, the most potent of which is farnesyl pyrophosphate (Oh et al., 2008). GPR92 is expressed in several tissues, most prominently in the CD8+ intraepithelial lymphocytes of the gastrointestinal tract (Kotarsky et al., 2006). Millipore's cloned human LPA5-expressing cell line is made in the Chem-1 host, which supports high levels of recombinant LPA5 expression on the cell surface and contains high levels of the promiscuous G protein Galpha15 to couple the receptor to the calcium signaling pathway. Thus, the cell line is an ideal tool for screening for agonists, antagonists and modulators at LPA5.