Immunohistochemistry and immunofluorescence
Immunohistochemical staining was performed on 3-μm paraffinized sections of renal tissue. Detection of CD68, CD4, CD8 and CD20 (Maxim-Bio), tryptase and CD1c (Abcam) was performed on paraffin sections by an immunohistochemical kit (DAKO, Santa Clara, USA) according to the manufacturer’s instructions. For immunofluorescence, the primary antibody against C3, IgA, IgG, and IgM (Gene-Bio) and Alexa Flour 488-conjugated secondary antibody (Jackson ImmunoRearch, West Grove, USA) were used. The numbers of each subset of interstitial inflammatory cells were counted under five equivalent high-power cortical fields (HPFs) (×400) and were expressed as the average number of cells per square millimeter [16]. The labeled cells were counted three times per high power field (HDF) to reduce counting errors, and the average number was recorded as the density of positive cells in this renal section.