Ra3 6b2

Kidney grafts were harvested at designated time points and either immediately submerged in 10% formalin for paraffin embedding or preserved in OCT; and 4 μm sections were used for HE staining. IHC and immunofluorescence (IF) staining were performed as published previously (38 (link)). The Abs used included anti-CD3(Cell Signaling Technology, D4V8L), B220 (BioLegend, RA3-6B2), and C4d (Novus Biologicals, 16D2). To test the ability of DSA produced from single-cell cultures to bind to alloantigens, supernatants from the highest 5 DSA-positive clones were pooled and used to bind allogeneic native Balb/c kidney sections for 24 hours at 4°C. Then, primary Ab anti-IgG (SouthernBiotech) was used to detect the IgG. Images were captured on a ZEISS Axiolab 5 microscope.

Kidney grafts were harvested at designated time points and either immediately submerged in 10% formalin for paraffin embedding or preserved in OCT; and 4 μm sections were used for HE staining. IHC and immunofluorescence (IF) staining were performed as published previously (38 (link)). The Abs used included anti-CD3(Cell Signaling Technology, D4V8L), B220 (BioLegend, RA3-6B2), and C4d (Novus Biologicals, 16D2). To test the ability of DSA produced from single-cell cultures to bind to alloantigens, supernatants from the highest 5 DSA-positive clones were pooled and used to bind allogeneic native Balb/c kidney sections for 24 hours at 4°C. Then, primary Ab anti-IgG (SouthernBiotech) was used to detect the IgG. Images were captured on a ZEISS Axiolab 5 microscope.