海西定
铁转运蛋白
单克隆抗体
内化
化学
抗体
生物素化
克隆(Java方法)
脂多糖
转铁蛋白
分子生物学
生物
生物化学
免疫学
细胞
炎症
基因
作者
Shinobu Sakamoto,Mika Kirinashizawa,Yumi Mohara,Yoshihiro Watanabe
摘要
ABSTRACT Hepcidin regulates the quantity of ferroportin (FPN) on cellular membrane. In our cell assay expressing ferroportin labeled with green fluorescence, FPN was internalized and degraded only after treatment with hepcidin-25, not hepcidin-22 or hepcidin-20, leading to accumulation of cellular iron. Thus we generated murine monoclonal antibodies (mAbs) against hepcidin-25, and then characterized and validated their functions. Among them, several mAbs showed a neutralizing activity that may prevent ferroportin internalization induced by hepcidin-25. To measure hepcidin level in various fluids, mAbs specific for human and rat hepcidin-25 were selected. As for rat, a sandwich ELISA developed using clone rHN1 as capture antibody and biotinylated clone mHW1 as a detection reagent had high sensitivity, allowing for the detection of 1-100 ng/mL of hepcidin-25. Rat hepcidin-25 level in plasma was measured at an average concentration of 63.0 ng/mL in healthy condition, and at 218.2 ng/mL after stimulation of lipopolysaccharide.
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