primary antibodies against vegfr2  (Cell Signaling Technology Inc)

Journal: bioRxiv

Article Title: Inference of VEGFR2 dimerization kinetics on the cell surface by integrating single-molecule imaging and mathematical modeling

doi: 10.1101/2025.06.03.657760

Figure Lengend Snippet: (A) Cartoon of FLR2 (full-length VEGFR2) and its ECTM truncation mutant, together with their HaloTag labeling. (B) Representative TIRFM image of Halo-FLR2 on the surface of CHO cells labeled at the single-molecule level with Halo-ligand conjugated to JF549, with cell outline in red. Halo JF549 labeling. Scale bar, 10 µm. The image is inverted for visual clarity. (C) Particle (i.e. labeled receptor complex) trajectories (with duration > 5 frames) from the cell shown in B, imaged for 30 s at 10 Hz. Trajectories are assigned random colors to help distinguish between them visually. Scale bar and red outline as in B. (D) Example of splitting and merging event from the boxed area in A-B. Scale bar, 10 µm. Orange arrow shows the particle when the two molecules are together, while cyan and magenta arrows show the two particles when separated. (E) Fraction of labeled receptor complexes, i.e., detected particles, in the indicated apparent oligomeric states, for FLR2 (blue) and ECTM (magenta). Circles show fractions from analyzing all imaged cells together. Error bars show the standard deviation of the fractions, estimated using Jackknife resampling. Note the cut in the y-axis. (F) Density of labeled receptor complexes per cell, shown as a distribution of individual cell measurements. Dashed vertical lines show mean values (0.157 for FLR2 and 0.156 for ECTM). (G) Density of labeled receptors per cell, shown as a distribution of individual cell values. Dashed vertical lines show mean values (0.164 for FLR2 and 0.171 for ECTM). (H, I) Apparent dimer dissociation (H) and association (I) rate constants. Circles and error bars as in E. (J) Diffusion coefficient distribution from all particles and all cells of each condition (FLR2 or ECTM). SMI dataset consisted of 85 (FLR2) and 94 (ECTM) cells, combined from 14 (FLR2) and 18 (ECTM) independent repeats.

Article Snippet: The samples were then incubated for 1 h with 1:1000 primary antibody against VEGFR2 (4B4; ThermoFisher, MA5-15556) in blocking buffer.

Techniques: Mutagenesis, Labeling, Standard Deviation, Diffusion-based Assay

Journal: bioRxiv

Article Title: Inference of VEGFR2 dimerization kinetics on the cell surface by integrating single-molecule imaging and mathematical modeling

doi: 10.1101/2025.06.03.657760

Figure Lengend Snippet: (A) Representative 2-color fixed-cell image of Halo-FLR2 in CHO cells labeled with Halo-ligand, together with an anti-phospho-VEGFR2 antibody, in the absence (top) or presence of 2 nM VEGF for 5 min. Cell outline is shown in red. The individual channel images are inverted for visual clarity. Scale bar, 10 µm. (B) Fraction of VEGFR2 detections (Halo-FLR2 in CHO cells or endogenous VEGFR2 in TIME cells) colocalized with the anti-phospho-VEGFR2 antibody, as a measure of VEFGR2 phosphorylation in the absence or presence of VEGF. Circles indicate individual cell measurements, with open circles indicating outliers. Red lines and shaded bars show mean and standard deviation, respectively, over group of cells. ***, p < 0.001, where -VEGF and +VEGF measurements were compared using a 2-sample t-test. The rise in Halo-FLR2 phosphorylation in CHO cells in response to VEGF is similar to that of endogenous VEGFR2 in TIME cells, indicating that Halo-FLR2 is able to bind VEGF and to get subsequently activated. Dataset consisted of 60, 69, 39 and 44 cells from 9, 10, 4 and 4 independent repeats for CHO -VEGF, CHO +VEGF, TIME -VEGF and TIME +VEGF, respectively.

Article Snippet: The samples were then incubated for 1 h with 1:1000 primary antibody against VEGFR2 (4B4; ThermoFisher, MA5-15556) in blocking buffer.

Techniques: Labeling, Phospho-proteomics, Standard Deviation

Journal: bioRxiv

Article Title: Inference of VEGFR2 dimerization kinetics on the cell surface by integrating single-molecule imaging and mathematical modeling

doi: 10.1101/2025.06.03.657760

Figure Lengend Snippet: (A) Representative 2-color fixed-cell image of FLR2 in CHO cells, labeled with Halo-ligand and anti-VEGFR2 antibody. Cell outline is shown in red. The individual channel images are inverted for visual clarity. Scale bar, 10 µm. (B, C) Fraction of antibody detections colocalized with Halo label detections vs the density of halo label detections for FLR2 (B) and ECTM (C) . Each dot represents one cell. Solid and dashed lines show straight line fit (solid line) and its 95% confidence interval (dashed lines). (D) Density of the full population of receptor complexes, as derived from the inverse of the slope of the line fits in B and C. Distribution stems from randomly sampling the distribution of slopes, assuming a Gaussian distribution with mean and standard deviation estimated by the least squares fit. Dashed vertical lines show mean values (1.33 for FLR2 and 1.44 for ECTM). (E) Fraction of labeled receptor complexes per cell in the SMI dataset, shown as a distribution of individual cell measurements. Dashed vertical lines show mean values (0.118 for both FLR2 and ECTM). 2-color fixed cell dataset consisted of 65 (FLRs) and 44 (ECTM) cells, combined from 10 (FLR2) and 6 (ECTM) independent repeats. SMI dataset used for E same as that in .

Article Snippet: The samples were then incubated for 1 h with 1:1000 primary antibody against VEGFR2 (4B4; ThermoFisher, MA5-15556) in blocking buffer.

Techniques: Labeling, Derivative Assay, Sampling, Standard Deviation

Journal: Discover Oncology

Article Title: Anticancer effect of the antirheumatic drug leflunomide on oral squamous cell carcinoma by the inhibition of tumor angiogenesis

doi: 10.1007/s12672-025-01763-5

Figure Lengend Snippet: Immunohistochemistry study of tumor specimens of syngeneic mice. a Representative microscopic images of the specimen of syngeneic mice. HE staining and IHC staining for DHODH and the tumor vascular endothelial markers integrin αv, VEGFR2, and CD31 of the tumors from the control (DMSO vehicle control) and LEF-treated (LEF-treated group) groups. Ten fields of view were taken for each specimen, and images of representative fields of view (sample number 4, 5 as control and 8, 9 as test in Fig. ) are shown on the left at low magnification and right at high magnification of the areas highlighted in white rectangle. Black bars in all panels indicated 500 µm. ( b ) Quantitative analysis of IHC positivity rates was conducted by calculating the ratio of the stained area of each marker, DHODH and vascular endothelial cells, to the horizontal projected area of the sections. This analysis was performed using BZ-X Analyzer software (BZ-H4A, Keyence Corporation, Osaka, Japan), with five sections randomly selected for each marker. Statistical analysis was performed using Student’s t-test. Values are expressed as means ± SEM of five sections (* p < 0.05)

Article Snippet: Immunostaining was performed using primary antibodies against vascular endothelial growth factor receptor 2 (VEGFR2) (rabbit mAb, 55B11; at 1: 100, Cell Signaling Technology, Danvers, MA, US), integrin alpha V rabbit mAb (ab179475; at 1: 500), CD31 mouse mAb (ab182981; at 1: 2000, Abcam Inc., Cambridge, MA, USA), DHODH rabbit pAb (14877-1-AP; at 1: 300, Proteintech, Rosemont, IL, USA).

Techniques: Immunohistochemistry, Staining, Control, Marker, Software