Quantification of Phospholipase C Activation and NFκB Signaling
Activation of phospholipase C was determined by quantification of the [3H]-inositol phosphates (InsP) [21 (link)]. NFκB-luciferase reporter gene assays were performed in HEK293T cells [52 (link)] and luminescence (1 s per well) was measured with a Mithras LB940 multilabel plate reader (Berthold Technologies). VEGF levels were measured in conditioned medium by ELISA using the Quantikine human-VEGF ELISA kit (R&D systems) [25 (link)]. In all VEGF ELISA assays, the lower detection limit was 15 pg/ml.
Heukers R., Fan T.S., de Wit R.H., van Senten J.R., De Groof T.W., Bebelman M.P., Lagerweij T., Vieira J., de Munnik S.M., Smits-de Vries L., van Offenbeek J., Rahbar A., van Hoorick D., Söderberg-Naucler C., Würdinger T., Leurs R., Siderius M., Vischer H.F, & Smit M.J. (2018). The constitutive activity of the virally encoded chemokine receptor US28 accelerates glioblastoma growth. Oncogene, 37(30), 4110-4121.
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