DiscussionThe monitoring of key signaling nodes, such as the MAP-Kinase ERK or PI3 Kinase/AKT, became a commonly used approach in investigations on signaling networks. These proteins represent either cellular readouts in a cascade of signaling events or are considered as the primary targets for compounds development. Therefore, the quantification of their phosphorylation status is of a great importance. For many years, the western-blot was the most popular technique to detect protein phosphorylation. More recently, assays based on the HTRF method, an advanced fluorescence-based technology, were developed to facilitate the research on phospho-proteins and more generally on signaling pathways.
Here we provide valuable results obtained with different HTRF assay on a large panel of cellular models, such as well-established cell lines grown in 2D or 3D, normal primary cells such as PBMC, or primary tumor and metastatsis patient-derived cancer. All together, these data show that the HTRF phospho-proteins platform can be easily implemented to investigate a large diversity of sample types, including primary cells.
Cruz T (1), Vallaghé J (2), Chouvet C (2), Morcillo V (2), Douzon S (2), Costy C (2), Oberleitner S (2), Trinquet E (2), Pannequin J (1), Charrier-Savournin F (2)
(1)Institut de génomique Fonctionnelle, CNRS UM5203, INSERM U661, Montpellier, France
(2)Cisbio Bioassays, Codolet, France