Journal: Neoplasia (New York, N.Y.)

Article Title: EGFR Regulates the Development and Microarchitecture of Intratumoral Angiogenic Vasculature Capable of Sustaining Cancer Cell Intravasation

doi: 10.1016/j.neo.2015.08.002

Figure Lengend Snippet: EGFR-mediated development of intravasation-sustaining vasculature involves the tumor cell-produced VEGF released by neutrophil MMP-9. A) Gene expression of VEGF and IL8 was determined by RT-qPCR in siCtrl or siEGFR HT-hi/diss cells. Data are fold difference over GAPDH levels (1.0) calculated from two independent experiments. (B) Microtumors were initiated from siCtrl and siEGFR HT-hi/diss cells. Individual siEGFR microtumors were treated daily with 10 ng purified neutrophil MMP-9 or equal volume of vehicle (10 μl). On day 5, soluble (PBS-extracted) VEGF and IL-8 were quantified in microtumor extracts by a capture ELISA. Data are percentage of control (100%) calculated from three independent experiments. The density of neutrophils was analyzed in two independent experiments by immunofluorescent staining of microtumors for MMP-9, a specific marker of MMP-9–loaded neutrophils. Image on the right depicts neutrophils isolated from the chick embryo and immunostained with mAb specific for chicken MMP-9 (pink). DAPI staining (light blue) highlights multilobed nuclei, characteristic of neutrophils. (C–G) Fraction of siEGFR microtumors was treated daily with 10 ng neutrophil MMP-9, 25 ng VEGF, or equal volume of vehicle (10 μl). On day 5, microtumors were analyzed for overall appearance of the vasculature (C), blood vessel density (D), vessel diameter (E), and tumor vasculature permeability (F and G) as described in Figures 3 and . Data are from three independent experiments employing eight embryos for each treatment condition (49 siCtrl tumors, 49 siEGFR tumors, 22 siEGFR tumors treated with neutrophil MMP-9, and 48 siEGFR tumors treated with VEGF). (H) Intravasation levels were quantified by Alu -qPCR and presented as percentage of control (100%) for each treatment condition from two independent experiments involving 14 siCtrl-embryos, 14 siEGFR-embryos, 14 siEGFR-embryos treated with neutrophil MMP-9, and 15 siEGFR-embryos treated with VEGF.

Article Snippet: The levels of human VEGF and human IL-8 were analyzed by capture ELISA kits from Peprotech (900-M10 and 900-M18, respectively), according to the manufacturer’s instructions.

Techniques: Produced, Gene Expression, Quantitative RT-PCR, Purification, Enzyme-linked Immunosorbent Assay, Control, Staining, Marker, Isolation, Permeability

Journal: Frontiers in Immunology

Article Title: Dengue Serotype Cross-Reactive, Anti-E Protein Antibodies Confound Specific Immune Memory for 1 Year after Infection

doi: 10.3389/fimmu.2014.00388

Figure Lengend Snippet: Pre-existing dengue-specific IgG antibodies are not indicative of the anamnestic IgG response 4–7 days after fever onset . (A) Plasma samples were sorted according to the UV-DENV-2 titer measured at <72 h and the values were compared with the E DENV-2 titers at the same time point and with UV-DENV-2 titer measured 4–7 days after onset of fever. (B) Plasma samples were sorted according to their UV-DENV-2 titer measured at 4–7 days after onset of fever and compared with E DENV-2 and E DENV-1 titers at the same time point. Patient IDs in red indicate secondary infections whereas patient IDs in black indicates primary infections. The classification primary versus secondary infection was made based on pre-existing IgG measured with the commercial indirect PanBio ELISA kit (see ) in samples collected <72 h after fever onset. *Indicates samples that were not available.

Article Snippet: The commercially available PanBio Indirect ELISA kit (Inverness Medical, Australia) was used to classify patients experiencing a secondary infection based on their IgG seropositivity in the sample collected within 72h after fever onset.

Techniques: Infection, Enzyme-linked Immunosorbent Assay