sheep anti hcd47 polyclonal antibodies  (R&D Systems)

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Relationship between tumor-associated macrophage subsets and CD47 expression in squamous cell carcinoma of the head and neck in the tumor microenvironment.

doi: 10.1038/labinvest.2016.70

Figure Lengend Snippet: Figure 2 (a) CD47 surface expression in 5 SCCHN cell lines HSC-2, HSC-3, HSC-4, CA9-22, and Gun-1. All 5 cells were expressed CD47, and HSC-3 was the highest. (b) Phagocytosis of CFSE-labeled HSC-3 cells (green) by induced M2 macrophages stained with CD68 (pink) in a culture slide (arrow). Immunofluorescent staining confirmed that cancer cells were engulfed within 3 h by macrophages induced with our protocol. (c) Phagocytosis of HSC-3 cells by macrophages visualized by May–Grunwald–Giemsa staining. (i) A smear of HSC-3 cell suspension. HSC-3 cells had a small and a relatively reddish nucleus. (ii) A smear of peripheral blood. White blood cells including monocytes had a strong blue nucleus. (iii) Co-culture of induced macrophages and HSC-3 for 4 h. Large macrophages engulfed HSC-3 with small nuclei in vesicles in cytoplasm of the macrophages. (d) Phagocytosis of CD47+ HSC-3 by induced M1 (black bar) and M2 (white bar) subsets in vitro. A polyclonal sheep anti-human CD47-blocking IgG was added to media at concentrations of 0, 0.5, and 2 μg/ml at the time of the co-culture with macrophages and the CD47+ SCCHN cell line HSC-3. Summary experiments were repeated 8 times. Whole actual cell counts for analyses were 8162 ± 695 of M1 and 7393 ± 667 of M2.

Article Snippet: Slides were incubated at 4 oC overnight with polyclonal sheep anti-human CD47 IgG (AF4670, R&D Systems; 5 μg/ml), monoclonal mouse anti-human CD163 mAb (clone 10D6; 1:200; Leica Biosystems, Nussloch, Germany), and monoclonal mouse antihuman CD68 mAb (clone PG-M1; ready-to-use; Dako).

Techniques: Expressing, Labeling, Staining, Suspension, Co-Culture Assay, In Vitro, Blocking Assay

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Relationship between tumor-associated macrophage subsets and CD47 expression in squamous cell carcinoma of the head and neck in the tumor microenvironment.

doi: 10.1038/labinvest.2016.70

Figure Lengend Snippet: Figure 3 (a) CD47 expression on HSC-3 was more effectively, but not completely, knocked down by CD47 siRNA than by negative siRNA. Gray filled line: stained with isotype control Ab; Green: CD47 staining of HSC-3 without any siRNA; Blue: CD47 expression with negative siRNA; Red: CD47 expression with CD47 siRNA. (b) Slight increases were observed in the phagocytosis of HSC-3 by the induced M1 subset (black bar, P = 0.08), but not by the induced M2 subset (white bar), with the knockdown of CD47 using siRNA in vitro. Summary experiments were repeated in triplicate. Whole actual cell counts for analyses were 8404 ± 1715 of M1 and 9683 ± 839 of M2.

Article Snippet: Slides were incubated at 4 oC overnight with polyclonal sheep anti-human CD47 IgG (AF4670, R&D Systems; 5 μg/ml), monoclonal mouse anti-human CD163 mAb (clone 10D6; 1:200; Leica Biosystems, Nussloch, Germany), and monoclonal mouse antihuman CD68 mAb (clone PG-M1; ready-to-use; Dako).

Techniques: Expressing, Staining, Control, Knockdown, In Vitro

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Relationship between tumor-associated macrophage subsets and CD47 expression in squamous cell carcinoma of the head and neck in the tumor microenvironment.

doi: 10.1038/labinvest.2016.70

Figure Lengend Snippet: Figure 4 Demonstrative IHC staining of CD47 (a–c), CD68 (d), and CD163 (e) in OSCC tissue magnified by × 400. CD47 staining was evaluated when the percentages of stained tumor cells in an entire lesion were o25% as (−). (a), 25–75% as ( ± ) (b), and 475% as (+) (c). (a) No stained cancer cells, but positive red blood cells. (b) The upper left tumor population was CD47−, whereas the upper right population was CD47+ in a membranous manner. (c) Most cancer cells were CD47+. More than 4 areas of a representative field adjacent to cancer cells were counted at ×400 magnification for CD68+

Article Snippet: Slides were incubated at 4 oC overnight with polyclonal sheep anti-human CD47 IgG (AF4670, R&D Systems; 5 μg/ml), monoclonal mouse anti-human CD163 mAb (clone 10D6; 1:200; Leica Biosystems, Nussloch, Germany), and monoclonal mouse antihuman CD68 mAb (clone PG-M1; ready-to-use; Dako).

Techniques: Immunohistochemistry, Staining

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Relationship between tumor-associated macrophage subsets and CD47 expression in squamous cell carcinoma of the head and neck in the tumor microenvironment.

doi: 10.1038/labinvest.2016.70

Figure Lengend Snippet: Figure 5 Survival analyses by the Kaplan–Meiermethod. (a) The OS of CD47+ cases ( ± and +cases) was significantly shorter (P = 0.015). (b) Increases in the infiltration of CD68+ macrophages in SCCHN tissue were associated with shorter OS (P = 0.035). (c, d) Overall (c, P = 0.025) and progression-free (d, P = 0.011) survivals correlated with the infiltration of CD163+ M2.

Article Snippet: Slides were incubated at 4 oC overnight with polyclonal sheep anti-human CD47 IgG (AF4670, R&D Systems; 5 μg/ml), monoclonal mouse anti-human CD163 mAb (clone 10D6; 1:200; Leica Biosystems, Nussloch, Germany), and monoclonal mouse antihuman CD68 mAb (clone PG-M1; ready-to-use; Dako).

Techniques: