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

Article Title: Nephronectin is Correlated with Poor Prognosis in Breast Cancer and Promotes Metastasis via its Integrin-Binding Motifs

doi: 10.1016/j.neo.2018.02.008

Figure Lengend Snippet: Uncropped images of immunoblots displayed in main figures. (A) Uncropped version of immunoblot displayed in main B . Membranes were incubated with anti-V5 (CST) and GAPDH (Abcam) antibodies prior to development. Inset shows cropped area shown in main figure. (B-E) Uncropped versions of immunoblots displayed in main A . The membranes were cut horizontally prior to incubation with antibodies for ALIX (CST), V5 (CST), GAPDH (Abcam), GM130 (BD Biosciences) and CHMP4B (Atlas Antibodies/Sigma).

Article Snippet: IF detection was performed using anti-CHMP4B (Atlas Antibodies/Sigma, Cat.: HPA051751, Lot No.: R67106, dilution 1:300, anti-V5 (CST, Cat.: 13202S, Lot No.: 2, dilution 1:150) and secondary antibodies; Goat anti-Mouse IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 647, (Thermo Fisher Scientific, Cat.: A32728, Lot No.: RJ243424, dilution 1:1000) and Goat anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Thermo Fisher Scientific, Cat.: A32731, Lot No.: RJ243417, dilution 1:1000).

Techniques: Western Blot, Incubation

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

Article Title: Nephronectin is Correlated with Poor Prognosis in Breast Cancer and Promotes Metastasis via its Integrin-Binding Motifs

doi: 10.1016/j.neo.2018.02.008

Figure Lengend Snippet: NPNT is localized in extracellular vesicles. (A) Western blot of isolated microvesicles and exosomes from 66cl4-cells expressing EV, NPNT wild-type, NPNT RGE, or NPNT RGE-AIA detected with anti-V5 antibodies (CST). ALIX and CHMP4B were used as markers for microvesicles and exosomes, GM130 as a negative control and GAPDH for normalization control. Whole-cell lysates of 66cl4-NPNT cells were included as control. The images are cropped to display only relevant bands. Full-length blots are shown in . IF showing colocalization between the exosomal marker CHMP4B and NPNT detected with anti-CHMP4B and NPNT antibodies (Abnova) and visualized with Alexa Fluor 647 and Alexa Fluor 488 secondary antibodies, respectively, of (B) lung samples from in vivo lung colonization assay and (C) MMTV-PyMT mammary tumor samples. Images are representative of a series from five mice. Nuclei were stained with Hoechst. -ab reflects a control with no primary antibody. Scale bar; 5 μm NPNT is localized in extracellular vesicles. (A) Western blot of isolated microvesicles and exosomes from 66cl4-cells expressing EV, NPNT wild-type, NPNT RGE, or NPNT RGE-AIA detected with anti-V5 antibodies (CST). ALIX and CHMP4B were used as markers for microvesicles and exosomes, GM130 as a negative control and GAPDH for normalization control. Whole-cell lysates of 66cl4-NPNT cells were included as control. The images are cropped to display only relevant bands. Full-length blots are shown in Supplementary Figure S3. IF showing colocalization between the exosomal marker CHMP4B and NPNT detected with anti-CHMP4B and NPNT antibodies (Abnova) and visualized with Alexa Fluor 647 and Alexa Fluor 488 secondary antibodies, respectively, of (B) lung samples from in vivo lung colonization assay and (C) MMTV-PyMT mammary tumor samples. Images are representative of a series from five mice. Nuclei were stained with Hoechst. -ab reflects a control with no primary antibody. Scale bar; 5 μm

Article Snippet: IF detection was performed using anti-CHMP4B (Atlas Antibodies/Sigma, Cat.: HPA051751, Lot No.: R67106, dilution 1:300, anti-V5 (CST, Cat.: 13202S, Lot No.: 2, dilution 1:150) and secondary antibodies; Goat anti-Mouse IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 647, (Thermo Fisher Scientific, Cat.: A32728, Lot No.: RJ243424, dilution 1:1000) and Goat anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Thermo Fisher Scientific, Cat.: A32731, Lot No.: RJ243417, dilution 1:1000).

Techniques: Western Blot, Isolation, Expressing, Negative Control, Marker, In Vivo, Staining

Journal: Nature Communications

Article Title: Exploiting induced pluripotent stem cell-derived macrophages to unravel host factors influencing Chlamydia trachomatis pathogenesis

doi: 10.1038/ncomms15013

Figure Lengend Snippet: Results are the average of three independent measurements±s.d. using the Luminex customised anti-human cytokine Milliplex kit. *Represents statistically significant different ( P <0.05) between uninfected and Chlamydia- infected as determined using two-way ANOVA.

Article Snippet: TNF-α, IL-1β, IL-6, IL-8 and IL-10 were measured using a Millipore customised Milliplex anti-human cytokine kit and data acquired on a Luminex FLEXMAP 3D system (Millipore).

Techniques: Luminex, Infection

Journal: Nature Communications

Article Title: Exploiting induced pluripotent stem cell-derived macrophages to unravel host factors influencing Chlamydia trachomatis pathogenesis

doi: 10.1038/ncomms15013

Figure Lengend Snippet: Pathways upregulated in human iPSdMs upon C. trachomatis infection as shown by proteomics.

Article Snippet: TNF-α, IL-1β, IL-6, IL-8 and IL-10 were measured using a Millipore customised Milliplex anti-human cytokine kit and data acquired on a Luminex FLEXMAP 3D system (Millipore).

Techniques: Infection, Activation Assay, Coagulation

Journal: Nature Communications

Article Title: Exploiting induced pluripotent stem cell-derived macrophages to unravel host factors influencing Chlamydia trachomatis pathogenesis

doi: 10.1038/ncomms15013

Figure Lengend Snippet: ( a , c ) Level of GFP-tagged C. trachomatis infection of human iPSdM CRISPR/Cas9 mutants after 48 h. Results are the averages from three independent measurements±s.d. using the Cellomics CellInsight NXT. ( b ) Production of cytokines from the IRF5 −/− mutant. Results are the average from three biological replicates assessed using the Luminex Multiplex and shown as fold change relative to expression of each cytokine in KOLF2 parent iPSdMs. *Represent statistically significant different ( P <0.05) between parent and mutants as determined using two-way ANOVA. Equal numbers of WT and mutant cells were seeded for each experiment.

Article Snippet: TNF-α, IL-1β, IL-6, IL-8 and IL-10 were measured using a Millipore customised Milliplex anti-human cytokine kit and data acquired on a Luminex FLEXMAP 3D system (Millipore).

Techniques: Infection, CRISPR, Mutagenesis, Luminex, Multiplex Assay, Expressing