Cd206 antibody

Hepatic tissue samples were obtained, formalin-fixed and paraffin-embedded. As in our previously published paper, haematoxylin and eosin (HE) staining (Jin et al., 2015 (link)) and Masson's trichrome staining (Li et al., 2016 (link)) were performed. Masson's trichrome stains hepatic cells in red and collagen in blue, which could help to clarify the degree of fibrosis. CD206 was a cell surface marker which could distinguish M2 macrophages from M1 macrophages. Immunohistochemistry for CD206 was performed to observe the M2 macrophages distribution in the liver tissue sections. The operation and analysis was performed according to our previously published protocol (Du et al., 2016 (link)). CD206 antibody was purchased from Proteintech Group, INC (Chicago, USA), catalog number 18704-1-AP, and its dilution was 1:200. The dilution of CD206 antibody was 1:200. The granuloma area and IOD of CD206 was measured by Imag-Pro Plus 6.0.

Proteins were extracted from the colonic tissues or murine BMDM using a standard extraction reagent supplemented with the protease inhibitor (KANGCHEN; Shanghai, China). The protein concentration was determined using a bicinchoninic acid protein assay kit (Beyotime Institute of Biotechnology, Haimen, China). Proteins were then separated using sodium dodecyl sulfate polyacrylamide gel electrophoresis and electro-transferred to nitrocellulose membranes as described previously^{68 (link)}, and incubated with a primary antibody overnight at 4 °C. The samples were then incubated with an IRDye800CW-conjugated secondary antibody (Rockland, Gilbertsville, PA, USA) for 1 h at 25 °C. The image was acquired with the Odyssey infrared imaging system (Li-Cor Biosciences, Lincoln, NE, USA). All of the immunoblotting experiments were repeated at least five times. The following primary antibodies were used: Beclin-1 antibody (1:500; Cell Signaling Technology, Danvers, MA, USA), light chain 3 (LC3) antibody (1:500; Novus Biologicals, Littleton, CO, USA), and p62 antibody (1:500; Cell Signaling Technology, Danvers, MA, USA), CD68 antibody (1:300; Proteintech Group Inc., Chicago, IL, USA), and CD206 antibody (1:500; Proteintech Group Inc., Chicago, IL, USA).