Immunohistochemistry was performed using frozen sections as described previously [25] (link), [26] (link). The following antibodies were used as primary antibodies: (1) polyclonal rabbit anti-type IV collagen antibody (Chemicon International, Inc., Temecula, CA); (2) polyclonal guinea pig anti-nephrin antibody (Fitzgerald, Concord, MA); (3) polyclonal rabbit anti-ZO-1 antibody (ZYMED Laboratories, Carlsbad, CA) and (4) monoclonal rat anti-CD31 antibody (Pharmingen, San Diego, CA). The glomerular accumulation of monocytes/macrophages was determined by immunohistochemistry using monoclonal rat anti-Mac-2 (lectin, galactoside-binding, soluble, 3) antibody (Cedarlane, Burlington, Ontario, Canada) as previously described [30] (link).
Double immunofluorescent staining was performed as previously described [10] (link), [11] (link). The following antibodies were used as primary antibodies: (1) polyclonal rabbit anti-phosphorylated NF-κB p65 (pNF-κB p65) antibody (Cell Signaling Technology, Danvers, MA); (2) monoclonal rat anti-CD34 antibody (Santa Cruz Biotechnology, CA).
Double immunofluorescent staining was performed as previously described [10] (link), [11] (link). The following antibodies were used as primary antibodies: (1) polyclonal rabbit anti-phosphorylated NF-κB p65 (pNF-κB p65) antibody (Cell Signaling Technology, Danvers, MA); (2) monoclonal rat anti-CD34 antibody (Santa Cruz Biotechnology, CA).