Vegf165

Manufactured by R&D Systems

Sourced in United States, Germany

VEGF165 is a recombinant human vascular endothelial growth factor protein. It is a member of the VEGF family and plays a role in angiogenesis, the formation of new blood vessels.

Automatically generated - may contain errors

Lab products found in correlation

Close spelling variants of this product

Recombinant human VEGF165 (36 citations) Human VEGF165 (10 citations) VEGF165 antibody (2 citations) Recombinant human VEGF (VEGF165) (2 citations) Carrier-free human VEGF165 (2 citations) Biotinylated human VEGF165 (2 citations)

64 protocols using vegf165

Efficient Generation of iPSC-derived Endothelial Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

ECs induction of PBMNCs-derived iPSCs were performed according to the previously
published protocol.^{29 (link)} In brief, iPSCs were passaged at 1:50–100 and cultured with mTeSR1 for four days.
Then, iPSCs culture medium were refreshed with BSA polyvinylalcohol essential lipids (BEL)
medium supplemented with Activin A (25 ng/mL, R&D), BMP4 (30 ng/mL, R&D), VEGF165
(50 ng/mL, R&D), and CHIR99021 (1.5 µM, Tocris Bioscience, Bristol, UK) for three days
to generate mesoderm cells. For vascular cells specification, mesoderm induction medium
were substituted by BEL medium supplemented with VEGF165 (50 ng/mL), SB431542 (10 µM,
Millipore, Billerica, MA, USA) for four days. To expand vascular ECs, cells were treated
with the same medium as vascular specification for 3–4 more days. Next, mature vascular
endothelial (VE) cells were purified by using CD31-dynabeads (Miltenyi Biotec, Bergisch
Gladbach, Germany). The purified VE cells were maintained and expanded in EC-serum-free
media (SFM, Gibco) containing VEGF165 (30 ng/mL), bFGF (30 ng/mL, R&D), and fetal
bovine serum (FBS, 1%, Gibco).

Zhou F., Zhao X., Liu X., Liu Y., Ma F., Liu B, & Yang J. (2020). Autologous correction in patient induced pluripotent stem cell-endothelial cells to identify a novel pathogenic mutation of hereditary hemorrhagic telangiectasia. Pulmonary Circulation, 10(4), 2045894019885357.

+ Open protocol

+ Expand

Quantitative VEGF Sandwich ELISA

Check if the same lab product or an alternative is used in the 5 most similar protocols

A monoclonal antibody specific for human VEGF had been pre-coated onto 96 well polystyrene microplate (VEGF¹⁶⁵, R&D System, Cat No: MAB3045-SP) at concentration 1000 pg/mL in a buffered protein base with preservatives; lyophilized. The test samples were applied (100μL/well) in duplicate. A standard curve was obtained, using 200 μL/well of VEGF¹⁶⁵ (R&D systems) at concentrations of 0, 31.3, 62.5, 125, 250, 500, 1000, 2000 pg/mL, in duplicate wells. Each well was aspirated and washed, and the process was repeated twice for a total of three washes. Each well was washed by washing buffer (400 μL) using a squirt bottle, manifold dispenser, or autowasher. A 200 μL of human VEGF conjugate was added to each well. Subsequently, each well was covered with a new adhesive strip and incubated for 2 hours at room temperature. After that, we repeated the aspiration/wash procedure and added 200 μL of substrate solution to each well. We then incubated each well for 25 minutes at room temperature, followed by adding 50 μL of stop solution (2N Sulfuric acid) to each well. We determined the optical density of each well within 30 minutes, using a microplate reader set to 450 nm.

Prodjohardjono A., Vidyanti A.N., Susianti N.A., S.u.d.a.r.m.a.n.t.a., Sutarni S, & Setyopranoto I. (2020). Higher level of acute serum VEGF and larger infarct volume are more frequently associated with post-stroke cognitive impairment. PLoS ONE, 15(10), e0239370.

+ Open protocol

+ Expand

Functionalized MWNT for VEGF Delivery

Check if the same lab product or an alternative is used in the 5 most similar protocols

The versatile features of MWNT are functionalized, such as the multipurpose innovative carriers for drug delivery and diagnostic applications via physicochemical treatment. In our study, it means the MWNT are purified and treated by plasma polymerization to obtain the ability to carry and release VEGF₁₆₅. Raw MWNT (Wako Pure Chemical Industries, Ltd, Osaka, Japan) was purchased and purified as previously reported.18 (link) VEGF₁₆₅ was purchased from R&D Systems, Inc. (Minneapolis, MN, USA). Recombinant human VEGF₁₆₅ was loaded into purified MWNT at a ratio of 1:10,000 (0.1 μg:1 mg) by solution sonication for 30 minutes. The suspensions were then freeze-dried for 48 hours. The VEGF₁₆₅-loaded MWNT were coated with PLGA film by plasma polymerization to prepare the VEGF₁₆₅-releasing device, as reported previously.19 The ratio of lactic acid to glycolic acid was 1:2. After plasma polymerization for 12 hours, the novel MWNT were characterized using high-resolution transmission electron microscopy (HRTEM). The amount of VEGF₁₆₅ loaded into MWNT was analyzed by human VEGF enzyme-linked immunosorbent assay (ELISA) kits (Quantikine® Colorimetric Sandwich ELISAs, R&D Systems), according to the manufacturer’s instructions.

Liu Z., Feng X., Wang H., Ma J., Liu W., Cui D., Gu Y, & Tang R. (2014). Carbon nanotubes as VEGF carriers to improve the early vascularization of porcine small intestinal submucosa in abdominal wall defect repair. International Journal of Nanomedicine, 9, 1275-1286.

+ Open protocol

+ Expand

Reagents and Methods for Cell Imaging

Check if the same lab product or an alternative is used in the 5 most similar protocols

Unless noted otherwise, MEM and all other cell culture reagents were purchased from Life Technologies. For [Ca²⁺]_i imaging studies, 35-mm dishes with glass coverslip windows were purchased from MatTek Corp. Fura-2 AM and Pluronic F127 were obtained from Life Technologies. CaCl₂ was from EMD Milllipore. ATP (disodium salt), PMA and all other chemicals, unless noted otherwise, were from Sigma. VEGF-165 and PlGF (placental growth factor) were from R & D Systems, Inc. VEGF-E, a product of recombinant orf virus, was from Angio-Proteiomie. VEGFRi; (VEGF receptor tyrosine kinase inhibitor; 4-[(4’-chloro-2’-fluoro)phenylamino]-6,7-dimethoxyquinazoline, a reasonably selective inhibitor of VEGFR-2 over VEGFR-1; IC50 = 100 nM and 2 uM, respectively) was purchased from EMD Millipore. PP2 (a selective inhibitor of Src family kinases) was from Enzo Life Sciences. U0126 (a selective inhibitor of MEK, a kinase known to directly phosphorylate ERK1 and 2) was from Promega Corp. CLA isomers were initially provided by Dr. Mark Cook and Dr. Manish Patankar. Subsequent isomers were purchased from Matreya LLC, with no difference in data between the two sources.

Boeldt D.S., Grummer M.A., Yi F., Magness R.R, & Bird I.M. (2015). Phosphorylation of Ser-279/282 and Tyr-265 Positions on Cx43 as Possible Mediators of VEGF-165 Inhibition of Pregnancy-Adapted Ca2+ Burst Function in Ovine Uterine Artery Endothelial Cells. Molecular and cellular endocrinology, 412, 73-84.

+ Open protocol

+ Expand

HUVEC Tube Formation Assay with Jarid2 and miR-130a

Check if the same lab product or an alternative is used in the 5 most similar protocols

Tube formation assays were performed using HUVEC cells as described elsewhere [34 (link)]. Briefly, for tube formation assays, HUVECS were transfected with Jarid2 constructs (0.25μg) using a nucleofector kit (Lonza), followed by plating 0.6 × 10⁵ cells on a 24-well plate coated with Low Growth Factor Matrigel (BD Biosciences, San Jose, CA) and supplemented with 50ng/ml of VEGF₁₆₅ (R&D Systems, MN) in serum-free media. The tubes were imaged at 10X magnification and quantified from 4 different fields. Co-transfection of Jarid2 and miR-130a (100nM) was performed as described above.

Singh B.N., Tahara N., Kawakami Y., Das S., Koyano-Nakagawa N., Gong W., Garry M.G, & Garry D.J. (2017). Etv2-miR-130a-Jarid2 cascade regulates vascular patterning during embryogenesis. PLoS ONE, 12(12), e0189010.

+ Open protocol

+ Expand

VEGF165 Binding Assay for Bevacizumab

Check if the same lab product or an alternative is used in the 5 most similar protocols

The immunoreactivity assay was conducted according to previous reported^{43 (link), 44 (link)}. Each well in ELISA plates (Corning Incorporated, USA) was coated with 5 μg/ml of recombinant human VEGF165 (R&D Systems) at 4 °C overnight, and then VEGF165 solution was adjusted to pH 9.6 by adding 15 mM Na₂CO₃ and 35 mM NaHCO₃. Thereafter, the wells were blocked with 100 μL 1% human serum albumin (HSA, Sigma) in 0.067 M PBS. After blocking, the wells were washed three times with 0.1% polysorbate 80 (Sigma) in 0.067 M PBS. For the following binding assay, 10 ng/ml of ^99mTc-MAG₃-bevacizumab was added to the wells with VEGF coated. All wells were allowed to incubate for 2 h at room temperature. Subsequently, unbound antibody was removed by washing the wells three times with 0.1% polysorbate 80 in 0.067 M PBS. Finally, the bound antibody was solubilized with 0.2 M NaOH and withdrawn for gamma counting. The percentage of binding was calculated as bound counts divided by total counts. Competition experiments were performed by adding an excess of unlabeled bevacizumab (1000 fold) to the wells one hour in advance before ^99mTc-MAG₃-bevacizumab was added into VEGF165 coated wells. Each group was conducted in triplicate.

Tan H., Zhou J., Yang X., Abudupataer M., Li X., Hu Y., Xiao J., Shi H, & Cheng D. (2017). 99mTc-labeled bevacizumab for detecting atherosclerotic plaque linked to plaque neovascularization and monitoring antiangiogenic effects of atorvastatin treatment in ApoE−/− mice. Scientific Reports, 7, 3504.

+ Open protocol

+ Expand

Cell Culture Reagents and Assay Protocols

Check if the same lab product or an alternative is used in the 5 most similar protocols

IL-1β, TNF, EGF, bFGF and VEGF 165 were purchased from R&D systems. Hydrocortisone, YN1, and DMEM without glucose, L-glutamine, sodium pyruvate powder were purchased from Sigma Aldrich. Fetal calf serum (FCS), gentamicin, fungizone, L-glutamine, MCBD 131, Opti-MEM and TRI-reagent were purchased from Thermo Fisher Scientific. Trypsin and EDTA were purchased from BioWhittaker. 3PO was purchased from Merck Millipore and dissolved in DMSO. HEPES buffer, Rotenone, Antimycin A, and 2-deoxyglucose were purchased from Agilent in the glycolytic rate assay starter kit.

Wik J.A., Lundbäck P., la Cour Poulsen L., Haraldsen G., Skålhegg B.S, & Hol J. (2020). 3PO inhibits inflammatory NFκB and stress-activated kinase signaling in primary human endothelial cells independently of its target PFKFB3. PLoS ONE, 15(3), e0229395.

+ Open protocol

+ Expand

Cytokine-Induced Metabolic Regulation

Check if the same lab product or an alternative is used in the 5 most similar protocols

Interleukin‐1β, EGF, bFGF, and VEGF 165 were from R&D systems (Minneapolis, MN, USA). Hydrocortisone, l‐glutamine, sodium pyruvate, glucose (45%), oligomycin, carbonyl cyanide‐4‐(trifluoromethoxy) phenylhydrazone (FCCP), and Dulbecco's Modified Eagle Medium (DMEM) without glucose (D5030) were from Sigma‐Aldrich (Milwaukee, WI, USA). FBS, gentamicin, fungizone, MCDB 131, and TRI reagent were from Thermo Fisher Scientific (Waltham, MA, USA). Trypsin and ethylenediaminetetraacetic acid tetrasodium salt dihydrate (EDTA) were from BioWhittaker (Walkersville, MD, USA). HEPES buffer was from Agilent (Santa Clara, CA, USA).

Wik J.A., Phung D., Kolan S., Haraldsen G., Skålhegg B.S, & Hol Fosse J. (2021). Inflammatory activation of endothelial cells increases glycolysis and oxygen consumption despite inhibiting cell proliferation. FEBS Open Bio, 11(6), 1719-1730.

+ Open protocol

+ Expand

Optimizing VEGF165 Dose for MSCs

Check if the same lab product or an alternative is used in the 5 most similar protocols

To establish the optimal concentration of VEGF₁₆₅ (R & D, USA), in our system, a dose/response curve was performed, using P3 MSCs obtained from both one control and one patient.
Each experiment was performed in triplicate and the optimal stimulation dose for VEGF was always assessed to be 50 ng/mL.

Cipriani P., Di Benedetto P., Capece D., Zazzeroni F., Liakouli V., Ruscitti P., Pantano I., Berardicurti O., Carubbi F., Alesse E, & Giacomelli R. (2014). Impaired Cav-1 expression in SSc mesenchymal cells upregulates VEGF signaling: a link between vascular involvement and fibrosis. Fibrogenesis & Tissue Repair, 7, 13.

+ Open protocol

+ Expand

Angiogenic Growth Factor Inhibitors

Check if the same lab product or an alternative is used in the 5 most similar protocols

PDGF-BB was purchased from Miltenyi Biotech (Bergisch Gladbach, Germany), VEGF-165 from R&D Systems (Minneapolis, MN, USA). InSolution™ Akt inhibitor IV, and PI3K inhibitor PKI-179 were obtained from Calbiochem, (San Diego, CA, USA). The Glucose Transporter Inhibitor IV, WZB117, was purchased from Merck Millipore (Darmstadt, Germany).

Moench R., Grimmig T., Kannen V., Tripathi S., Faber M., Moll E.M., Chandraker A., Lissner R., Germer C.T., Waaga-Gasser A.M, & Gasser M. (2016). Exclusive inhibition of PI3K/Akt/mTOR signaling is not sufficient to prevent PDGF-mediated effects on glycolysis and proliferation in colorectal cancer. Oncotarget, 7(42), 68749-68767.

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!