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目的 针对人体肠道样本获取受限的现状，建立并验证适用于标准化小鼠模型的肠道固有层免疫细胞亚群（T细胞、B细胞、固有免疫细胞）多色流式检测方案，以支持小鼠模型肠道黏膜免疫应答水平的系统评估。方法 基于细胞膜表面分子标志物，包括分化簇（Cluster of differentiation, CD）45、CD3、CD4、CD19、CD38、CD45、CD69、CD185、程序性死亡受体1(Programmed cell death 1, PD-1)、趋化因子受体5(Chemokine receptor 5, CCR5)等，制定两套15色流式检测方案，分别用于评价肠道固有层T细胞与固有免疫细胞亚群以及用于评估固有层B淋巴细胞亚群。采用C57BL/6小鼠制备肠道固有层单个核淋巴细胞悬液，验证方案在评价小鼠肠道粘膜固有层免疫应答中的适用性。结果 应用所建立的流式方案，成功鉴定了包括活化T细胞、调节性T细胞、巨噬、NK、B、浆细胞等多个关键肠道固有层免疫细胞亚群，并通过检测刺激条件下T细胞亚群功能状态的特异性变化，实现对细胞免疫功能的精准评估。结论 本研究成功建立的两套多色流式方案适用于小鼠肠道固有层单个核淋巴细胞悬液样本，为监测小鼠模型肠道免疫微环境状态提供了可靠且高效的检测方案。
Abstract：Objective Given the limited availability of human intestinal samples, this study aimed to establish and validate a multicolor flow cytometry method for the detection of intestinal lamina propria immune cell subsets, including T cells, B cells, and innate immune cells, in standardized mouse model, thereby supporting systematic evaluation of intestinal mucosal immune responses. Methods Based on cell surface markers, including cluster of differentiation(CD)45, CD3, CD4, CD19, CD38, CD69, CD185, programmed cell death protein 1(PD-1), and chemokine receptor 5(CCR5), two 15-color flow cytometry panels were designed: one for the evaluation of lamina propria T-cell and innate immune cell subsets, and the other for the assessment of lamina propria B-cell subsets. Mononuclear lymphocyte suspensions from the intestinal lamina propria were prepared from C57 BL/6 mice, and the applicability of the two panels for assessing mucosal immune responses in the murine intestinal lamina propria was validated. Results Using the established flow cytometry panels, multiple key immune cell subsets in the intestinal lamina propria were successfully identified, including activated T cells, regulatory T cells, macrophages, natural killer(NK) cells, B cells, and plasma cells. In addition, by detecting specific functional changes in T-cell subsets under stimulatory conditions, the assay enabled precise evaluation of cellular immune function. Conclusion The two multicolor flow cytometry panels established in this study are suitable for mononuclear lymphocyte suspensions from the murine intestinal lamina propria and provide a reliable and efficient method for monitoring the intestinal immune microenvironment in mouse models.
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基本信息:
DOI：10.13242/j.cnki.bingduxuebao.250395
中图分类号:R392
引用信息:
[1]王倩莹,杜欣然,王硕,等.用于小鼠模型肠道固有层免疫细胞亚群多色流式检测方法的建立[J].病毒学报,2026,42(03):812-822.DOI:10.13242/j.cnki.bingduxuebao.250395.
基金信息:
国家重点研发计划（项目号：2024YFC2309502）,题目：HIV/MPXV共感染及痘苗疫苗接种人群免疫特征多组学研究~~
2026-05-07
2026-05-07
2026-05-07