Human Phospho-Axl (Y779) Antibody

Detection of Human Axl by Western Blot

Clinical drug responses were evaluated in the patient-derived primary culture GBM cells. (a) Most of the primary culture cancer cells were CD133 (green) and SOX2 (red), highly expressed using flow cytometry analysis and immunofluorescence assay. Hoechst33342-labeled nuclei. Bar = 50 μm. MFI: mean fluorescent intensity. (b) Flow cytometry analysis and immunofluorescence assay in GBM primary cultured spheroid cells to detect the high expression of CD133 (green) and SOX2 (red) cultured with serum-free medium. Hoechst33342-labeled nuclei. Bar = 50 μm. MFI: mean fluorescent intensity. (c) Primary culture cancer cells, treated with (Z)-BP and temozolomide (active form MTIC was used), presented relatively low IC50, compared to BCNU. Additionally, dosing (Z)-BP in IC30 concentration could reduce the required amount of TMZ for determining the synergistic effect on eliminating primary culture gliomas. (d) Using (Z)-BP (200 or 400 μM) to expose the primary culture cells could reduce the level of MGMT, in comparison to vehicle control and BCNU (800 μM). Further, receptor tyrosine kinases including EGFR, phosphorylated EGFR, and Axl were also downregulated. Upstream mTOR activity was inactivated (p-mTOR) by (Z)-BP, and this could mutually affect Axl and EGFR signaling and then decrease the protein level of PD-L1. beta -Actin was used as an internal control. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/35646111), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Phospho-Axl (Y779) by Western Blot

The combination of sitravatinib with abemaciclib or palbociclib is highly toxic against TNBC cells. (a) Chemical structure of sitravatinib (Sitra). (b) Immunoblot was performed on cell lines treated for 24 h with Abe (2 μm), Palbo (5 μm), and/or Sitra (2 μm). Protein levels were determined for phospho-AXL, phosho-MET, and phosho-MERTK. (c) The clonogenic assay showing that the combination of Abe or Palbo with Sitra significantly decreased the colony formation capacity of TNBC cells. Representative images of stained colonies. (d) Combination index (CI) values for the combinations of sitravatinib or merestinib with CDK4/6 inhibitor abemaciclib using different doses. Circles represent experimentally determined CI values using the Chou–Talalay method. The colors (orange and blue) represent the fixed ratio mixtures. (e,f) Overview of the toxicity and synergy scores of the drug combinations for TNBC lines. The heatmaps show the level of toxicity (e) and Bliss number (f) for the cell lines tested in this study. Average values of toxicity (e) or Bliss number (f) for cells treated with sitravatinib (S) at varying doses (S0 = No Drug, S1 = 1 μm, S2 = 2 μm, and S3 = 3 μm) in combination with either abemaciclib (A) at varying doses (A0 = No Drug, A1 = 1 μm, A2 =2 μm, A3 = 3 μm, and A4 = 4 μm) or palbociclib at varying doses (P0 = No Drug, P1 = 1 μm, P2 = 2 μm, P3 = 3 μm, and P4 = 4 μm). (g) Shown is the caspase-3/7 activity measured upon 24 h of drug treatments. The data are presented as mean ± SEM from three independent experiments, expressed as ratios to untreated control values, with associated p values as indicated (One-way ANOVA with Dunnett’s multiple comparisons test analysis). Abe: abemaciclib; Palbo: palbociclib. The original western blot figures can be found in File S1. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/38927958), licensed under a CC-BY license. Not internally tested by R&D Systems.