Ferritins are spherical iron storage proteins within cells, composed of 24 subunits of two types, heavy-chain ferritin (HFn) and light-chain ferritin. Ferritins auto-assemble naturally into hollow nanocages with an outer diameter of 12 nm and an interior cavity 8 nm in diameter. Since the intrinsic tumor-targeting property of human HFn was first reported in 2012, HFn has been extensively explored for tumor-targeted delivery of anticancer drugs and diagnostic molecules, including radioisotopes and fluorophores, as well as inorganic nanoparticles (NPs) and chemotherapeutic drugs. This protocol provides four detailed procedures describing how to load four types of cargoes within HFn nanocages that are capable of accurately controlling cargo loading: synthesis of inorganic metal nanoparticles within the cavity of a wild-type human HFn nanocage (Procedure 1, requires ~5 h); loading of doxorubicin into the cavity of a wild-type human HFn nanocage (Procedure 2, requires ~3 d); loading Gd³⁺ into the cavity of a genetically engineered human HFn nanocage (Procedure 3, requires ~20 h); and loading ⁶⁴Cu²⁺ radioisotope into the cavity of a genetically engineered human HFn nanocage (Procedure 4, requires ~3 h). Subsequent use of these HFn-based formulations is advantageous as they have intrinsic tumor-targeting capability and lack immunogenicity. Human HFn generated as described in this protocol can therefore be used to deliver therapeutic drugs and diagnostic signals as multifunctional nanomedicines.

铁蛋白是细胞内的球形铁储存蛋白，由重链铁蛋白 (HFn) 和轻链铁蛋白两种类型的 24 个亚基组成。铁蛋白自然地自动组装成外径为 12 nm、内部空腔直径为 8 nm 的中空纳米笼。自 2012 年首次报道人类 HFn 的内在肿瘤靶向特性以来，HFn 已被广泛探索用于肿瘤靶向递送抗癌药物和诊断分子，包括放射性同位素和荧光团，以及无机纳米颗粒 (NP) 和化疗药物。该协议提供了四个详细的程序，描述了如何在 HFn 纳米笼内装载四种类型的货物，这些货物能够准确控制货物装载：在野生型人类 HFn 纳米笼的腔内合成无机金属纳米颗粒（程序 1，需要约 5 小时） ）；将阿霉素装入野生型人 HFn 纳米笼的空腔中（程序 2，需要约 3 天）；将 Gd ³⁺加载到基因工程人类 HFn 纳米笼的空腔中（程序 3，需要约 20 小时）；将⁶⁴ Cu ²⁺放射性同位素加载到基因工程人类 HFn 纳米笼的空腔中（程序 4，需要约 3 小时）。这些基于 HFn 的制剂的后续使用是有利的，因为它们具有内在的肿瘤靶向能力并且缺乏免疫原性。因此，按本协议所述生成的人类 HFn 可用于作为多功能纳米药物传递治疗药物和诊断信号​​。