Journal: Biochimica et biophysica acta

Article Title: Interaction of the human prostacyclin receptor and the NHERF4 family member intestinal and kidney enriched PDZ protein (IKEPP).

doi: 10.1016/j.bbamcr.2012.07.015

Figure Lengend Snippet: Fig. 8. Effect of IKEPP on hIP agonist-induced migration and tube formation in 1° HUVECs. Migration (panels A and B) and tube formation (panels C and D) by primary (1°) HUVECs were analyzed at 0 h and 12 h in following incubation with drug vehicle, VEGF or cicaprost, as indicated, in either non-transfected cells (panels A and C) or in cells transfected with siRNACTRL or siRNAIKEPP (panels B and D), where immunoblot analysis confirmed specific disruption of IKEPP expression (panel E). Bar charts represent mean fold increases (±S.E.M.; n=3) in either wound closure or tube length in the presence of vehicle, VEGF or cicaprost, as indicated, at 12 h. The asterisks indicate either significant agonist-induced increases in migration (panels A and B) or significant agonist-induced increases in tube length (panels C and D) in comparison to vehicle-treated cells where **, and *** indicate Pb0.01, and Pb0.001, respectively, for post hoc Dunnett's multiple comparison t-test analysis.

Article Snippet: Rabbit anti-IKEPP antibody was from Abcam®; mouse monoclonal anti-hemaglutinin (HA) 101R antibody was from Cambridge Biosciences; goat anti-IKEPP, normal rabbit IgG, horseradish peroxidase (HRP)-conjugated goat anti-rabbit, rabbit antigoat and goat anti-mouse secondary antibodies were from Santa Cruz; rat monoclonal anti-HA 3F10-HRP-conjugated antibody was from Roche; rabbit anti-PhosphoSer antibody was from Invitrogen (61– 8100); recombinant human vascular endothelial growth factor (VEGF) 165 (293-VE/CF) was from R&D Systems; mouse monoclonal antiFLAG and anti-FLAG-HRP-conjugated M2 antibody, protein Gsepharose and protein A-sepharose were from Sigma-Aldrich; antiHDJ-2 (DNAJ protein) was from Neomarkers; AlexaFluor488 goat anti-rabbit, and AlexaFluor594 goat anti-mouse antibodies were from Molecular Probes.

Techniques: Migration, Incubation, Transfection, Western Blot, Disruption, Expressing, Comparison

Journal: The Oncologist

Article Title: A Phase I Open‐Label Clinical Trial Evaluating the Therapeutic Vaccine hVEGF26 –104/ RFASE in Patients with Advanced Solid Malignancies

doi: 10.1002/onco.13576

Figure Lengend Snippet:

Article Snippet: VEGF serological responses: Anti‐VEGF antibody titers were measured in serum, frozen the day the material was received, and stored at −80°C until analysis, using an in‐house developed ELISA. Microplates were coated with 100 μL recombinant hVEGF 165 (1 μg/mL; BioLegend, San Diego). After washing, the plates were blocked with 200 μL 4% horse serum (Sigma‐Aldrich, St. Louis, MO). Hereafter, the plates were incubated with 100 μL 1:30 diluted serum. Horseradish peroxidase (HRP)–conjugated rabbit antihuman immunoglobulin G antibodies (1:8,000 dilution; Sigma‐Aldrich, St. Louis) were applied to detect bound antibodies in the microplate wells. In the presence of chromogenic substrate TMB (R&D Systems, Abingdon, UK) color was developed by the enzymatic reaction of HRP. Absorbance was measured using a BioTek Synergy HT plate reader at an OD of 450 nm. If the OD was above a predetermined cut‐off (mean + 3 SDs of all patient serum baseline OD levels), a relevant antibody response was suspected and a dilution series was performed. The antibody titer was defined as the 10 logarithm of the highest dilution which resulted in a signal above the predetermined cut‐off. A similar ELISA was performed on all samples to measure antibodies recognizing VEGF 26–104 . , .

Techniques: Neutralization, Protein Concentration, Enzyme-linked Immunosorbent Assay, Recombinant, Incubation, Release Assay, Isolation, Centrifugation, Cell Culture, Modification, Negative Control, Flow Cytometry, FCAP Assay, Software