Rabbit polyclonal anti sap polyclonal antibody

Formalin-fixed paraffin-embedded sections, cut at 3 μm, were deparaffinized, and antigen retrieval was performed at 99° C. The sections were reacted with rabbit polyclonal anti-SAP polyclonal antibody (1:400; ThermoFisher, Waltham, MA) followed by a Rhodamine red X-conjugated goat anti-rabbit secondary which was solid-phase adsorbed to ensure minimal cross-reaction with human IgG (1:100; Jackson ImmunoResearch Laboratories, West Grove, PA). Each case was run with positive and negative controls. The stain was evaluated by standard immunofluorescence microscopy. It was judged to be positive if there was positive granular capillary loop staining in the glomeruli, and negative if there was no capillary loop staining in the glomeruli. Colocalization of IgG and SAP in the glomerular basement membranes was examined by confocal microscopy using a Zeiss LSM 880 confocal laser scanning microscope (Zeiss Microscopy, Jena, Germany). For this analysis, polyclonal (fluorescein isothiocyanate–conjugated) rabbit anti-human IgG (1:40; Agilent, Santa Clara, CA) was reacted with the heat retrieved tissue following the staining for SAP as described above. Negative controls were performed to ensure antibody specificity by omitting primary antibodies. Four cases of MGMID were tested for the presence of HP1BP3 (1:100; Thermo Fisher, Waltham, MA) by immunoperoxidase staining.