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2 protocols using il 10

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Immunohistochemical Analysis of IL-10 and Foxp3

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Formalin-fixed, embedded histopathological slides were obtained from the Department of Pathology. Briefly, the slides were dewaxed and rehydrated with ethanol, and autoclaved at 121°C for 10 min. The sections were treated with 3% hydrogen peroxide for 30 min, permeabilized with 1% Triton X-100 for 15 min and incubated with normal goat serum for 15 minutes. Primary antibodies of IL-10 (GeneTex, Irvine, CA, USA) and Foxp3 (Santa Cruz Biotechnology, Santa Cruz, CA, USA) were incubated with sections at 4°C overnight before biotinylated secondary-antibodies were applied. All sections were then stained with DAB (Beyotime Biotechnology, Shanghai, China). Nuclei were counterstained with Meyer’s haematoxylin (Beyotime Biotechnology, Shanghai, China).
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2

Cellular Responses to Iron Compounds

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Ferric sulfate and ferrous sulfate heptahydrate were purchased from Energy Chemical Technology Co., Ltd. (Shanghai, China). Ferric citrate was purchased from Sigma‐Aldrich (St. Louis, MO). Fetal bovine serum (FBS) was purchased from Invitrogen (Carlsbad, CA). Dulbecco's Modified Eagle's Medium (DMEM) was obtained from Sigma (Yuzhong District, Chongqing, China). Antibodies against p21 and C646, a p300/CREB‐binding protein (CBP) inhibitor, were purchased from Abcam Trading Co. Ltd. (Shanghai, China). N‐acetyl‐l‐cysteine (NAC), dihydroethidium (DHE), and a ROS Detection Kit were purchased from Beyotime Company (Jiangsu, China). Prussian Blue staining solution was acquired from Leagene Biological Technology Co. Ltd. (Beijing, China). Antibodies against p53, acetylated p53, CD86, and CD206 were from Cell Signaling Technology (Danvers, MA), and IL‐1, IL‐10, TGF‐β, and TNF‐α were purchased from GeneTex (San Antonio, TX). p21, p300, CBP, p65, phosphorylated p65, and glyceraldehyde 3‐phosphate dehydrogenase (GAPDH) were purchased from Abcam (Cambridge, MA). FITC‐CD86 and APC‐CD206 were purchased from Thermo Fisher Scientific (Waltham, MA).
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