Liu Yongxiang, Jin Huan, Huo Bitao, Xia Xiaojun, Liu Jinyun
State Key Laboratory of Oncology in South China, Guangdong Provincial Clinical Research Center for Cancer, Sun Yat-sen University Cancer Center, Guangzhou, P.R. China; Guangzhou National Laboratory, Guangzhou International Bio-Island, Guangzhou, P.R. China.
State Key Laboratory of Oncology in South China, Guangdong Provincial Clinical Research Center for Cancer, Sun Yat-sen University Cancer Center, Guangzhou, P.R. China.
STAR Protoc. 2025 Aug 9;6(3):104024. doi: 10.1016/j.xpro.2025.104024.
Multiple metabolic pathways and metabolites are involved in innate immune memory induction of macrophages; however, protocols for in vitro-trained immunity assays induced by metabolites in mouse macrophages are limited. Here, we present a protocol for measuring endogenous 24(S),25-epoxycholesterol and inducing innate immune memory in mouse macrophages. We describe steps for sample preparation, measurement of 24(S),25-epoxycholesterol, and establishment of an in vitro-trained immunity model. We then detail procedures for assays measuring cytokine concentration and for assay for transposase-accessible chromatin using sequencing (ATAC-seq). For complete details on the use and execution of this protocol, please refer to Liu et al..
多种代谢途径和代谢产物参与巨噬细胞的天然免疫记忆诱导；然而，由代谢产物在小鼠巨噬细胞中诱导的体外训练免疫测定方案有限。在此，我们提出一种用于测量内源性24(S),25-环氧胆固醇并在小鼠巨噬细胞中诱导天然免疫记忆的方案。我们描述了样品制备、24(S),25-环氧胆固醇测量以及建立体外训练免疫模型的步骤。然后，我们详细介绍了测量细胞因子浓度的测定程序以及使用测序法（ATAC-seq）进行转座酶可及染色质测定的程序。有关本方案使用和执行的完整详细信息，请参考Liu等人的研究。
