Potter Timothy M, Neun Barry W, Rodriguez Jamie C, Ilinskaya Anna N, Dobrovolskaia Marina A
Cancer Research Technology Program, Nanotechnology Characterization Laboratory, Leidos Biomedical Research, Inc., Frederick National Laboratory for Cancer Research, P.O. Box B, Frederick, MD, 21702, USA.
Methods Mol Biol. 2018;1682:173-187. doi: 10.1007/978-1-4939-7352-1_15.
Cytokines, chemokines, and interferons are released by the immune cells in response to cellular stress, damage and/or pathogens, and are widely used as biomarkers of inflammation. Certain levels of cytokines are needed to stimulate an immune response in applications such as vaccines or immunotherapy where immune stimulation is desired. However, undesirable elevation of cytokine levels, as may occur in response to a drug or a device, may lead to severe side effects such as systemic inflammatory response syndrome or cytokine storm. Therefore, preclinical evaluation of a test material's propensity to cause cytokine secretion by healthy immune cells is an important parameter for establishing its safety profile. Herein, we describe in vitro methods for analysis of cytokines, chemokines, and type II interferon in whole blood cultures derived from healthy donor volunteers. First, whole blood is incubated with controls and tested nanomaterials for 24 h. Then, culture supernatants are analyzed by ELISA to detect IL-1β, TNFα, IL-8, and IFNγ. The culture supernatants can also be analyzed for the presence of other biomarkers secreted by the immune cells. Such testing would require additional assays not covered in this chapter and/or optimization of the test procedure to include relevant positive controls and/or cell types.
细胞因子、趋化因子和干扰素由免疫细胞在应对细胞应激、损伤和/或病原体时释放，并且被广泛用作炎症的生物标志物。在诸如疫苗或免疫疗法等期望免疫刺激的应用中，需要一定水平的细胞因子来刺激免疫反应。然而，细胞因子水平的不良升高，如可能因药物或器械而发生的情况，可能会导致严重的副作用，如全身炎症反应综合征或细胞因子风暴。因此，对测试材料引发健康免疫细胞分泌细胞因子倾向的临床前评估是确定其安全性概况的一个重要参数。在此，我们描述了用于分析来自健康供体志愿者的全血培养物中细胞因子、趋化因子和II型干扰素的体外方法。首先，将全血与对照和测试纳米材料一起孵育24小时。然后，通过酶联免疫吸附测定法（ELISA）分析培养上清液以检测白细胞介素-1β（IL-1β）、肿瘤坏死因子α（TNFα）、白细胞介素-8（IL-8）和干扰素γ（IFNγ）。培养上清液也可以分析免疫细胞分泌的其他生物标志物的存在情况。此类测试将需要本章未涵盖的额外测定方法和/或优化测试程序以纳入相关阳性对照和/或细胞类型。
