Rabbit anti vegf antibody

Recombinant mouse IL-25 (rmIL-25, Cat#: 587306) was brought from Biolegend, recombinant human IL-25 (rhIL-25, Cat#: 8134-IL) was purchased from R&D systems and prepared following instructions. Rabbit anti-CD31 antibody (Cat#: GB11063-2) was obtained from Servicebio. Goat anti-IL-17RB antibody (Cat#: AF1040) was obtained from R&D systems. Rabbit anti-VEGF antibody (Cat#: ab52917) and rabbit anti-β-catenin antibody (Cat#: ab32572) was from Abcam. Mouse anti-GAPDH antibody (Cat#: 60004-1-Ig) and rabbit anti-IL-17RB antibody (Cat#: 20673-1-AP) were obtained from Proteintech. Rabbit anti-phospho-Akt antibody (Cat#: 4060T), rabbit anti-Akt antibody (Cat#: 4091T), rabbit anti-phospho-ERK 1/2 antibody (Cat#: 4370S) and rabbit anti-ERK 1/2 antibody (Cat#: 4695T) were purchased from Cell Signaling Technology (CST). Glucose and streptozotocin (STZ, Cat#: S0130) were purchased from Sigma-Aldrich; and pLenti-CMV-IL-25-GFP-puro lentiviral plasmid (Lenti-IL-25-GFP, Cat#: PPL02165-4a) was obtained from Public Protein/Plasmid Library (PPL). The lentiviral packaging plasmid pMD2.G (Cat#: 12259) and psPAX2 (Cat#: 12260) were obtained from AddGene. Basement membrane matrix (Cat#: 354234) was obtained from Corning.

Cells were harvested and lysed in lysis buffer (50 mM Tris–HCl, 150 mM NaCl, 1% Non-idet P-40, 0.5% sodium deoxycholate, 0.1% SDS, and 1 mM phenylmethylsulfonyl fluoride). A Bio-Rad protein assay kit was used to determine protein concentrations, with bovine serum albumin acting as a reference. Samples containing 30 μg of protein were put on 10% SDS-polyacrylamide gels and electrophoretically transferred to nitrocellulose membranes. Membranes were incubated with specific primary antibodies including rabbit anti-VEGF antibody (Abcam, 1:1,000), rabbit anti-MYB antibody (Abcam, 1:400) and rabbit anti-β-actin antibody (Abcam, 1:2,000), then membrans were incubated with horseradish peroxidase-conjugated secondary antibody (Sigma, 1:10,000). Proteins were evaluated using a SuperSignal West Pico chemiluminescence kit (Thermo Scientific, United States).