goat polyclonal anti vegf 164  (R&D Systems)

Journal: The Keio Journal of Medicine

Article Title: Intravenous Regeneration-associated Cell Transplantation Enhances Tissue Recovery in Mice with Acute Ischemic Stroke

doi: 10.2302/kjm.2024-0005-oa

Figure Lengend Snippet: Fig. 3. Effect of RACs on vascular endothelial growth factor (VEGF) expression in peri-infarct area. Injection of both low (1 × 104 /50 µL) and high RACs (1 × 105 /50 µL) on day 3 after MCAO resulted in a significant increase in VEGF- positive cells in the peri-infarct area compared to PBS (*P < 0.05, Kruskal–Wallis test) [PBS: n=15, RACs (1 × 104 /50 µL): n=9, and RACs (1 × 105 /50 µL): n=10].

Article Snippet: Fixed sections were incubated in 5 mM hydrogen peroxide for 10 min before being exposed to 5% normal goat serum for an additional 10 min. Overnight incubation of the sections was carried out with polyclonal anti-goat VEGF (AF493NA; R&D Systems, Minneapolis, MN, USA) at a 50-fold dilution and monoclonal anti-rat IL-10 (ab33471; Abcam, Cambridge, UK) at a 200-fold dilution in a humidified chamber maintained at 4 °C.

Techniques: Expressing, Injection

Journal: Nephrology, dialysis, transplantation : official publication of the European Dialysis and Transplant Association - European Renal Association

Article Title: Chronic hypoxia exacerbates diabetic glomerulosclerosis through mesangiolysis and podocyte injury in db/db mice.

doi: 10.1093/ndt/gfaa074

Figure Lengend Snippet: FIGURE 2: Urinalysis of the effects of hypoxia in db/db mice. (A–C) Comparison of (A) urinary albumin (n ¼ 6–8), (B) urinary PCX (n ¼ 6–9) and (C) urinary VEGF-A (n ¼ 6–8) levels between hypoxic (filled square) and age-matched normoxic (open square) mice. Vertical columns and bars depict mean 6 SD. (D and E) Photomicrographs of IF staining of urinary PCX in samples from db/db mice collected af- ter 1 week of hypoxia. (D) Negative PCX staining in sediments obtained with conventional centrifugation. (E) Strong granular PCX stain- ing in the bottom fraction after the subsequent UC of the supernatants obtained with conventional centrifugation. Bars ¼ 20 lm. *P < 0.05, **P < 0.01, ***P < 0.001.

Article Snippet: Similarly, goat anti-mouse VEGF polyclonal antibody (AF-493-NA; R&D Systems), rabbit anti-transforming growth factor-b1 (TGF-b1) monoclonal antibody (ab215715; Abcam) and mouse anti-eNOS monoclonal antibody (ab76198; Abcam) were used.

Techniques: Comparison, Staining, Centrifugation

Journal: Nephrology, dialysis, transplantation : official publication of the European Dialysis and Transplant Association - European Renal Association

Article Title: Chronic hypoxia exacerbates diabetic glomerulosclerosis through mesangiolysis and podocyte injury in db/db mice.

doi: 10.1093/ndt/gfaa074

Figure Lengend Snippet: FIGURE 4: Representative glomerular immunohistological stainings of (A) CD34-positive endothelial cells, (C) WT-1-positive podocytes, (E) VEGF-positive cells, (G) F4/80-positive macrophages (indicated by arrow), (I) TGF-b1and (K) eNOS (K) in 12- and 24-week-old normoxic and hypoxic db/db mice. The graphs on the right show the comparison of positive area densities (%) of (B) CD34, (D) WT-1, (F) VEGF, (J) TGF-b1and (L) eNOS and (H) the average number of F4/80-positive macrophages per glomerulus among normoxic (open circle) and hypoxic (filled circle) 12-week-old mice and normoxic (open triangle) and hypoxic (filled triangle) 24-week-old mice. Each symbol represents an individual. Horizontal lines and boxes are the same as those in Figure 3. *P < 0.05, **P < 0.01, ***P < 0.005 and ****P < 0.001.

Article Snippet: Similarly, goat anti-mouse VEGF polyclonal antibody (AF-493-NA; R&D Systems), rabbit anti-transforming growth factor-b1 (TGF-b1) monoclonal antibody (ab215715; Abcam) and mouse anti-eNOS monoclonal antibody (ab76198; Abcam) were used.

Techniques: Comparison

Journal: Nephrology, dialysis, transplantation : official publication of the European Dialysis and Transplant Association - European Renal Association

Article Title: Chronic hypoxia exacerbates diabetic glomerulosclerosis through mesangiolysis and podocyte injury in db/db mice.

doi: 10.1093/ndt/gfaa074

Figure Lengend Snippet: FIGURE 5: Comparison of relative mRNA levels within LCM-iso- lated glomeruli encoding (A) VEGF-A, (B) VEGFR-2, (C) eNOS, (D) TGF-b1, (E) MCP-1and (F) PAI-1 between 24-week-old nor- moxic (open triangle) and hypoxic (filled triangle) db/db mice. Each symbol represents an individual. Horizontal lines and boxes are the same as those in Figure 3. *P < 0.05 and **P < 0.01.

Article Snippet: Similarly, goat anti-mouse VEGF polyclonal antibody (AF-493-NA; R&D Systems), rabbit anti-transforming growth factor-b1 (TGF-b1) monoclonal antibody (ab215715; Abcam) and mouse anti-eNOS monoclonal antibody (ab76198; Abcam) were used.

Techniques: Comparison

Journal: Cancer research

Article Title: Radiation-Induced Enhancement of Antitumor T-cell Immunity by VEGF-Targeted 4–1BB Costimulation

doi: 10.1158/0008-5472.CAN-16-2105

Figure Lengend Snippet: Radiation potentiates VEGF-4–1BB aptamer conjugate-mediated antitumor immunity in the VEGFlow 4T07 tumor model. A, VEGF expression in 4T1 and 4T07 tumors. 4T1 or 4T07 tumor cells were injected subcutaneously into Balb/c mice and when tumors reached about 150 mm3, they were resected and stained for VEGF expression as described in Materials and Methods. B, Homing of VEGF aptamer to VEGF-expressing tumors. Balb/c mice were implanted with 4T07 tumors in the opposite flanks. When tumors reached an average diameter of 75 mm3, the tumors at the right flank were irradiated (XRT) with either 2 or 20 Gy and 7 days later 32P-labeled VEGF (VEGF) or scrambled aptamer were administered via the tail vein. Twenty-four hours later mice were sacrificed, tumors excised, and radioactivity was determined by scintillation counting. The difference between the 2 and 20 Gy irradiated groups injected with the VEGF conjugate and any of the other groups was statistically significant (P < 0.01) C, 4T07 tumor cells were implanted subcutaneously in Balb/c mice and irradiated when tumor reached an average of 75 mm3. Seven days later, mice were treated with VEGF-4–1BB conjugate or with a mixture of VEGF and 4–1BB aptamers administered via the tail vein three times 2 days apart starting 6 days postirradiation. D, 4T07 tumor cells were implanted subcutaneously in Balb/c mice and irradiated when tumor reached an average of 75 mm3. Six days later, mice were treated with 100 pmol (1×) of VEGF-4–1BB conjugate, mixture of VEGF and 4–1BB aptamers, or with 800 pmol (8×) of an agonistic 4–1BB antibody or isotype control (IgG) three times 2 days apart. Aptamer was administered via the tail vein, whereas antibody was administered intraperitoneally.

Article Snippet: Nonspecific immunoreactivity in slide-mounted tissue sections was blocked with Serum-Blocking Reagent D (R&D Systems), and incubated with goat polyclonal anti-mouse VEGF at 1:20 (R&D Systems) at 4°C overnight, washed with PBS, and incubated with biotinylated anti-goat secondary antibody (R&D Systems) for 40 minutes, followed by high-sensitivity streptavidin (HSS)-horseradish peroxidase (HRP; R&D Systems) for 30 minutes.

Techniques: Expressing, Injection, Staining, Irradiation, Labeling, Radioactivity, Control

Journal: Cancer research

Article Title: Radiation-Induced Enhancement of Antitumor T-cell Immunity by VEGF-Targeted 4–1BB Costimulation

doi: 10.1158/0008-5472.CAN-16-2105

Figure Lengend Snippet: Combination of radiation with 4-1BB antibody, but not with VEGF-4–1BB aptamer conjugate, elicits nonspecific immune responses and inflammation. Balb/c mice were injected subcutaneously with 4T07 tumor cells and when they reached an average diameter of 75 mm3, tumors were irradiated. One hundred pmol (1×) of VEGF-4–1BB aptamer conjugate or 800 pmol (8×) of either 4–1BB antibody or IgG isotype control was administered 6 days later via the tail vein three times 2 days apart. Mice were sacrificed 5 days following the last administration. Liver (A) and spleen (B) weight. Percentage of CD8+ cells in the liver (C) and spleen (D). E and F, Serum IFNγ (E) and percentage of intracellular granzyme B-expressing CD8+ cells (F). Tissue sections from the lung (G) and liver (H) were stained with hematoxylin and eosin and visualized by light microscopy. Arrows, H&E staining inflammatory foci in the liver treated with 100 pmol (1×) of 4–1BB antibody. I, ALT and AST levels in the circulation 14 days after irradiation (3 mice/group). NS, nonsignificant.

Article Snippet: Nonspecific immunoreactivity in slide-mounted tissue sections was blocked with Serum-Blocking Reagent D (R&D Systems), and incubated with goat polyclonal anti-mouse VEGF at 1:20 (R&D Systems) at 4°C overnight, washed with PBS, and incubated with biotinylated anti-goat secondary antibody (R&D Systems) for 40 minutes, followed by high-sensitivity streptavidin (HSS)-horseradish peroxidase (HRP; R&D Systems) for 30 minutes.

Techniques: Injection, Irradiation, Control, Expressing, Staining, Light Microscopy

Journal: Cancer research

Article Title: Radiation-Induced Enhancement of Antitumor T-cell Immunity by VEGF-Targeted 4–1BB Costimulation

doi: 10.1158/0008-5472.CAN-16-2105

Figure Lengend Snippet: Inhibition of nonirradiated lung metastasis by local radiation of subcutaneously implanted tumors and treatment with VEGF-4–1BB aptamer conjugates. 4T1 tumor cells were implanted subcutaneously in the right flank of Balb/c mice, and 8 days later mice were injected intravenously with 4T1 cells. Five days after intravenous injections when the subcutaneously growing tumors reached an average size of 75 mm3, they were irradiated with 12 Gy and 3 days later treated with VEGF-4–1BB aptamer conjugates or with a mixture of VEGF and 4–1BB aptamers three times 2 days apart. Five days before irradiation, 4T1 tumor cells were injected intravenously to establish métastasés in the lung. Mice were sacrificed when mice in the control group showed signs of morbidity (21 days post-intravenous tumor administration) and tumor burden was determined. A, Volume of subcutaneously implanted tumor at day of sacrifice. B, Time course of subcutaneous tumor growth. C, Lung weights measured when mice were sacrificed.

Article Snippet: Nonspecific immunoreactivity in slide-mounted tissue sections was blocked with Serum-Blocking Reagent D (R&D Systems), and incubated with goat polyclonal anti-mouse VEGF at 1:20 (R&D Systems) at 4°C overnight, washed with PBS, and incubated with biotinylated anti-goat secondary antibody (R&D Systems) for 40 minutes, followed by high-sensitivity streptavidin (HSS)-horseradish peroxidase (HRP; R&D Systems) for 30 minutes.

Techniques: Inhibition, Injection, Irradiation, Control

Journal: Cancer research

Article Title: Radiation-Induced Enhancement of Antitumor T-cell Immunity by VEGF-Targeted 4–1BB Costimulation

doi: 10.1158/0008-5472.CAN-16-2105

Figure Lengend Snippet: Inhibition of irradiated and nonirradiated tumors in mice treated with PD-1 antibody, CTLA-4 antibody, and VEGF-4–1BB aptamer conjugates. 4T1 tumor cells were implanted subcutaneously in the right and left flanks of Balb/c mice at days 0 and 8, respectively. At day 13 when the tumors in the right flank reached an average size of 75 mm3, they were irradiated once with 12 Gy, and 3 days later treated as shown three times 2 days apart. Tumor volume at day 19 postirradiation is shown for the irradiated tumor, right flank (RF; A) and for nonirradiated tumor, left flank (LF; seven or eight mice per group; B). Time course of tumor growth is shown in Supplementary Fig. S4. Statistical analysis: A, differences between untreated group and groups treated by radiation, CTLA-4 antibody, PD-1 antibody, CTLA-4 + PD-1 antibody, or VEGF-4–1BB conjugate were significant (P < 0.05), whereas no significant differences were seen among the treated groups. Combination with radiation was significant for all treatments, PD-1, P = 0.0075; CTLA-4, P = 0.0191; PD-1 + CTLA-4, P = 0.0164; VEGF-4–1BB, P = 0.0071. B, Differences between untreated or radiated group and groups treated with CTLA-4 antibody, PD-1 antibody, CTLA-4 + PD-1 antibody, and VEGF-4–1BB conjugate were significant (P < 0.05). Differences between groups treated with radiation and PD-1 antibody or with PD-1 + CTLA-4 antibody and groups treated with antibody alone was not significant. Differences were significant between CTLA-4 antibody versus radiation + CTLA-4 antibody, P = 0.0265, and VEGF-4–1BB aptamer conjugate and radiation + VEGF-4–1BB aptamer conjugate groups, P = 0.0412. C, Survival of mice treated as described above. Mice were sacrificed when either tumor reached 1,000 mm3. Difference between untreated group and either radiation, PD-1 antibody, radiation + PD-1 antibody, CTLA-4 antibody, and VEGF-4–1BB aptamer conjugate-treated groups showed a trend but did not reach statistical significance. Differences between radiation + CTLA-4 antibody, PD-1 + CTLA-4 antibodies with or without radiation, and radiation + VEGF-4–1BB aptamer conjugate-treated groups and rest of the treatment groups was significant. Differences between radiation + VEGF-4–1BB aptamer conjugates or with PD-1 + CTLA-4 antibodies groups and PD-1 + CTLA-4 antibodies or radiation + CTLA-4 antibody-treated groups exhibited a trend but did not reach statistical significance.

Article Snippet: Nonspecific immunoreactivity in slide-mounted tissue sections was blocked with Serum-Blocking Reagent D (R&D Systems), and incubated with goat polyclonal anti-mouse VEGF at 1:20 (R&D Systems) at 4°C overnight, washed with PBS, and incubated with biotinylated anti-goat secondary antibody (R&D Systems) for 40 minutes, followed by high-sensitivity streptavidin (HSS)-horseradish peroxidase (HRP; R&D Systems) for 30 minutes.

Techniques: Inhibition, Irradiation

Journal: Cancer research

Article Title: Radiation-Induced Enhancement of Antitumor T-cell Immunity by VEGF-Targeted 4–1BB Costimulation

doi: 10.1158/0008-5472.CAN-16-2105

Figure Lengend Snippet: Hematoxylin and eosin staining of sections from small intestine. A, Magnification, ×10; B, Magnification, ×40. Mice were treated as described in Fig. 2. Arrows, increased lymphocytic infiltrates in mice treated with CTLA-4 antibody, CTLA-4 + PD-1 antibodies with or without radiation, compared with untreated mice or mice treated with VEGF-4–1BB aptamer conjugate with or without radiation. Contours indicate areas of dense leukocytic infiltrates that appear to have deformed or displaced the normal crypt architecture seen in mice treated with CTLA-4 or PD-1 + CTLA-4 antibodies with or without radiation (see also Supplementary Fig. S5).

Article Snippet: Nonspecific immunoreactivity in slide-mounted tissue sections was blocked with Serum-Blocking Reagent D (R&D Systems), and incubated with goat polyclonal anti-mouse VEGF at 1:20 (R&D Systems) at 4°C overnight, washed with PBS, and incubated with biotinylated anti-goat secondary antibody (R&D Systems) for 40 minutes, followed by high-sensitivity streptavidin (HSS)-horseradish peroxidase (HRP; R&D Systems) for 30 minutes.

Techniques: Staining

Journal: Cancer research

Article Title: Radiation-Induced Enhancement of Antitumor T-cell Immunity by VEGF-Targeted 4–1BB Costimulation

doi: 10.1158/0008-5472.CAN-16-2105

Figure Lengend Snippet: Tumor infiltration of immune cell subsets. Subcutaneously implanted 75 to 100 mm3 4T1 tumor-bearing mice were irradiated and/or treated with VEGF-4–1BB aptamer conjugates 3 days postirradiation, three times 2 days apart, and 2 days after the last treatment tumors were excised, homogenized, and analyzed for immune cell subsets by flow cytometry or for viability as described in Materials and Methods (Treg, CD3+CD4+CD25+FOXP3+; Ml macrophages, CD11Bhigh F4/80int).

Article Snippet: Nonspecific immunoreactivity in slide-mounted tissue sections was blocked with Serum-Blocking Reagent D (R&D Systems), and incubated with goat polyclonal anti-mouse VEGF at 1:20 (R&D Systems) at 4°C overnight, washed with PBS, and incubated with biotinylated anti-goat secondary antibody (R&D Systems) for 40 minutes, followed by high-sensitivity streptavidin (HSS)-horseradish peroxidase (HRP; R&D Systems) for 30 minutes.

Techniques: Irradiation, Flow Cytometry

Journal: Journal of pharmacological sciences

Article Title: Lymphangiogenesis induced by vascular endothelial growth factor receptor 1 signaling contributes to the progression of endometriosis in mice.

doi: 10.1016/j.jphs.2020.05.003

Figure Lengend Snippet: Fig. 4. The mRNA expression levels of Vegfc and Vegfd in cultured macrophages and fibroblasts. (A) The effects of PlGF (0 and 10 ng/ml) on the levels of the mRNAs encoding VEGFR1, VEGF-C, and VEGF-D in cultured bone marrow-derived macrophages from WT and TK-/- mice. Data are expressed as the mean ± SEM (n ¼ 4e6 mice per group). *P < 0.05. (B) The effects of PlGF (0 and 1 ng/ml) on the levels of the mRNAs encoding VEGFR1, VEGF-C, and VEGF-D in L929 cells in the presence of an anti-VEGFR1 antibody (10 ng/ml) or IgG. Data are expressed as the mean ± SEM (n ¼ 4e6 mice per group). *P < 0.05.

Article Snippet: The sections were incubated with the following primary antibodies at 4 C overnight: a rabbit anti-mouse VEGFR1 polyclonal antibody (1:200; Abcam), a goat anti-mouse VEGFR1 polyclonal antibody (1:200; Santa Cruz Biotechnology, Santa Cruz, CA, USA), a rabbit anti-LYVE-1 polyclonal antibody (1:200; Abcam), a goat anti-LYVE-1 polyclonal antibody (1:200; R&D Systems Inc., Minneapolis, MN, USA), a rabbit anti-VEGF-C polyclonal antibody (1:200; Abcam), a goat anti-VEGFD polyclonal antibody (1:40; R&D Systems Inc.), a rat anti-mouse CD11b monoclonal antibody (1:200; BD Biosciences, San Jose, CA, USA), a rabbit anti-mouse S100A4 polyclonal antibody (1:200; Abcam), or a goat anti-mouse S100A4 polyclonal antibody (1:100; OriGene Technologies, Rockville, MD, USA).

Techniques: Expressing, Cell Culture, Derivative Assay