1 / 1

Neutralizing Activity of Antibodies Against VEGF, IL-8, ENA-78, and MMP9 in Cell Culture

This study explores the neutralizing efficacy of specific antibodies against VEGF, IL-8, ENA-78, and MMP9 in cell culture experiments. Human umbilical vein endothelial cells (HUVEC) and granulocytes were treated with recombinant cytokines and corresponding antibodies. We observed strong inhibition of target activities, evaluated through proliferation assays and ELISA for myeloperoxidase secretion. The methodology included using polyclonal and monoclonal antibodies at varying concentrations, demonstrating significant impacts on cell responses crucial for understanding inflammatory pathways.

felcia
Télécharger la présentation

Neutralizing Activity of Antibodies Against VEGF, IL-8, ENA-78, and MMP9 in Cell Culture

An Image/Link below is provided (as is) to download presentation Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. Content is provided to you AS IS for your information and personal use only. Download presentation by click this link. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server. During download, if you can't get a presentation, the file might be deleted by the publisher.

E N D

Presentation Transcript

  1. (a) (b) (c) (d) MMP9 + anti MMP9 MMP9 + Isotype Supplementary Figure 5: Antibodies for VEGF, IL-8, ENA-78 and MMP9 used in cell culture experiments (figure 4e) were tested for their neutralizing activity. A strong and almost complete inhibition was found for all tested antibodies [(a) VEGF; (b) IL-8; (c) ENA-78; (d) MMP9]. Supplementary Figure 5 Methods (a) Human umbilical vein endothelial cells (HUVEC) were treated with 5 ng/ml of recombinant human VEGF together with 2 µg/ml of polyclonal goat anti human VEGF antibodies (AF-293-NA, R&D Systems, USA) or isotype control. 48 hours after treatment a proliferation-assay (EZ4U, Biomedica, Vienna Austria) was performed according to the manufacturer's instructions. (b,c) Human granulocytes were obtained by lysis of red blood cells in whole blood of 4 healthy probands followed by selective separation of CD66 positive cells using a MACS MicroBead Kit (Miltenyi Biotec, Germany). Granulocytes were stimulated with 5 ng/ml recombinant human IL-8 or 5 ng/ml ENA-78. For inhibition 2 µg/ml monoclonal mouse anti human IL-8 antibodies (AF-293-NA, R&D Systems, USA), 2 µg/ml goat anti human ENA-78 antibodies (AB-254-PB, R&D Systems, USA) or isotype control antibodies were added. Myeloperoxidase secretion was measured by ELISA (DMYE00, R&D Systems, USA). (d) For MMP9 inhibition an enzyme assay for MMP-activity was used. A specific MMP-substrate was incubated (Bachem, Basel, Switzerland) for 1 hour with 5 ng/ml recombinant MMP9 (R&D) together with 2 µg/ml MMP9 antibody or isotype control. The emitted fluorescence was measured with a fluorometer (Fluostar Optima, BMG Labtech, Vienna, Austria).

More Related