Feraheme (FH) nanoparticles (NPs) have been used extensively for treatment of iron anemia (due to their slow release of ionic iron in acidic environments). In addition, injected FH NPs are internalized by monocytes and function as MRI biomarkers for the pathological accumulation of monocytes in disease. We have recently expanded these applications by radiolabeling FH NPs for positron emission tomography (PET) or single-photon emission computed tomography (SPECT) imaging using a heat-induced radiolabeling (HIR) strategy. Imaging FH NPs using PET/SPECT has important advantages over MRI due to lower iron doses and improved quantitation of tissue NP concentrations. HIR of FH NPs leaves the physical and biological properties of the NPs unchanged and allows researchers to build on the extensive knowledge obtained about the pharmacokinetic and safety aspects of FH NPs. In this protocol, we present the step-by-step procedures for heat (120 °C)-induced bonding of three widely employed radiocations (⁸⁹Zr⁴⁺ or ⁶⁴Cu²⁺ for PET, and ¹¹¹In³⁺ for SPECT) to FH NPs using a chelateless radiocation surface adsorption (RSA) approach. In addition, we describe the conversion of FH carboxyl groups into amines and their reaction with an N-hydroxysuccinimide (NHS) of a Cy5.5 fluorophore. This yields Cy5.5-FH, a fluorescent FH that enables the cells internalizing Cy5.5-FH to be examined using flow cytometry. Finally, we describe procedures for in vivo and ex vivo uptake of Cy5.5-FH by monocytes and for in vivo microPET/CT imaging of HIR-FH NPs. Synthesis of HIR-FH requires experience with working with radioactive cations and can be completed within <4 h. Synthesis of Cy5.5-FH NPs takes ∼17 h.

中文翻译：

---

Feraheme纳米粒子的热诱导放射性标记和荧光标记，用于PET / SPECT成像和流式细胞术。

Feraheme（FH）纳米颗粒（NPs）已广泛用于治疗铁性贫血（由于它们在酸性环境中缓慢释放离子铁）。此外，注射的FH NPs被单核细胞内在化，并作为MRI生物标记物用于疾病中单核细胞的病理性蓄积。我们最近通过使用热诱导放射性标记（HIR）策略对FH NPs进行放射性标记来进行正电子发射断层扫描（PET）或单光子发射计算机断层扫描（SPECT）成像，从而扩展了这些应用。使用PET / SPECT对FH NP成像比MRI具有重要优势，这是因为铁剂量较低且组织NP浓度的定量改进。FH NP的HIR使NP的物理和生物学特性保持不变，并使研究人员可以在获得的有关FH NP的药代动力学和安全性方面的广泛知识的基础上继续发展。在此协议中，我们介绍了加热（120°C）诱导的三种广泛使用的放射性阳离子（使用无螯合的放射性阳离子表面吸附（RSA）方法将FZ NPs转换为PET的⁸⁹ Zr ⁴⁺或⁶⁴ Cu ²⁺和SPECT的¹¹¹ In ³⁺。此外，我们描述了FH羧基向胺的转化及其与Cy5.5荧光团的N-羟基琥珀酰亚胺（NHS）的反应。这产生了Cy5.5-FH，一种荧光FH，它使使用Cy5.5-FH内在化的细胞能够使用流式细胞仪进行检查。最后，我们描述了单核细胞体内和体外摄取Cy5.5-FH以及HIR-FH NPs的体内microPET / CT成像的程序。HIR-FH的合成需要使用放射性阳离子的经验，并且可以在不到4小时内完成。Cy5.5-FH NP的合成需要约17小时。