Endocytic Capacity in Ocrl and Clcn5 Mice

PTs endocytic capacity of Ocrl and Clcn5 mice was examined by measuring β-lactoglobulin and dextran uptake. β-lactoglobulin was tagged with Cy5 using TM2 Ab labeling kit (Amersham) in accordance with the manufacturer’s instructions. Fifteen minutes after tail-vein injection of Cy5-β-lactoglobulin (1 mg/kg B.W., L3908, Sigma) or 30 min after injection of 10 kDa Alexa 647-dextran (6 mg/kg B.W.; D22914, Thermo Fisher Scientific), mice were anesthetized and their kidneys were harvested and processed for confocal microscopy. The endocytic capacity of Ocrl and Clcn5 mPTCs was examined by measuring albumin and dextran uptake as described previously (44 (link)). Briefly mPTCs were incubated at 37°C with 100 μg ml⁻¹ Alexa488-BSA (A13100, Thermo Fisher Scientific) or 250 μg ml⁻¹ Alexa 647-dextran diluted in medium without FBS supplementation, for 15 and 30 min, respectively. After washing, the cells were fixed in 4% PFA and processed for confocal microscopy.