Immunology Division, Walter and Eliza Hall Institute of Medical Research, Parkville, VIC, Australia.
Department of Medical Biology, University of Melbourne, Parkville, VIC, Australia.
Methods Mol Biol. 2024;2826:167-187. doi: 10.1007/978-1-0716-3950-4_13.
Division tracking dyes like Cell Trace Violet (CTV) enable the quantification of cell proliferation, division, and survival kinetics of human naïve B cell responses in vitro. Human naïve B cells exhibit distinct responses to different stimuli, with CpG and anti-Ig inducing a T cell-independent (TI) response, while CD40L and IL-21 promote a T cell-dependent (TD) response that induces isotype switching and differentiation into antibody-secreting cells (ASCs). Both stimulation methods yield valuable insights into the intrinsic programming of B cell health within individuals, making them useful for clinical investigations. For instance, quantitative analysis from these B cell populations could reveal biologically meaningful measurements such as the average number of division rounds and the time to cells' fate. Here, we describe a novel in vitro culture setup for CTV-labelled human naïve B cells and a method for obtaining precise time-based data on proliferation, division-linked isotype switching, and differentiation.
分裂追踪染料，如 CellTrace Violet（CTV），可用于量化体外人类初始 B 细胞反应的细胞增殖、分裂和存活动力学。人类初始 B 细胞对不同刺激表现出不同的反应，CpG 和抗 Ig 诱导非依赖 T 细胞（TI）反应，而 CD40L 和 IL-21 促进依赖 T 细胞（TD）反应，诱导同种型转换和分化为抗体分泌细胞（ASC）。这两种刺激方法都深入了解了个体内 B 细胞健康的内在编程，使其成为临床研究的有用工具。例如，这些 B 细胞群体的定量分析可以揭示具有生物学意义的测量结果，例如平均分裂轮数和细胞命运的时间。在这里，我们描述了一种新的体外 CTV 标记人类初始 B 细胞培养方案，以及一种获取增殖、与分裂相关的同种型转换和分化的精确基于时间的数据的方法。
