Figure 3

Binding of CHC in kidneys of brain-dead pigs during cold storage. Porcine kidneys were perfused by gravity with UW supplemented with 500 μg/mL CHC, with ¹¹¹In labeled CHC added to trace the conjugate. Fractions of the flow-through from the renal vein were collected during infusion of the conjugate (A; red line), and following incubation for 3 h on ice, fractions of the saline used to remove any unbound conjugate were also collected and analyzed in the gamma counter (A; black line). The amount of bound CHC was measured in the kidney tissue (B) resulting in 0.13 ± 0.02 mg/g binding to the cortex and 0.12 ± 0.01 mg/g binding to the medulla. In total, the kidneys contained 0.13 ± 0.01 mg/g (n = 4). In addition, kidneys (n = 2) were filled with a higher amount of tracer and imaged with Single-photon emission computed tomography (SPECT)/computed tomography (CT) (C; left panel, activity gradient in blue–red representing low–high activity). After washing out unbound CHC, the kidneys were imaged again (C; right panel) to assess any variation in binding to different regions. The binding was further evaluated by autoradiography. Regions were identified macroscopically on frozen sections (D; left panel), and higher amounts were detectable in the cortex and outer medulla compared to the inner medulla (D; right panel). Frozen sections were stained with avidin-FITC (green) to detect CHC in the vessels stained for von Willebrand factor (VWF in red: E; scale bar = 100 μm).