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Quantifying Human Naïve B Cell Proliferation Kinetics and Differentiation in Controlled In Vitro Cell Culture
Anthony J Farchione1,2, HoChan Cheon1,2, Philip D Hodgkin1,2
1Immunology Division, Walter and Eliza Hall Institute of Medical Research, Parkville, VIC, Australia.
Insights
Cell Trace Violet (CTV) dyes quantify human B cell proliferation and survival in vitro. This study presents a novel culture method for precise, time-based analysis of B cell responses to different stimuli.
Area of Science:
- Immunology
- Cell Biology
Background:
- Division tracking dyes like Cell Trace Violet (CTV) are crucial for quantifying human naïve B cell proliferation, division, and survival kinetics in vitro.
- Human naïve B cells display distinct T cell-independent (TI) and T cell-dependent (TD) responses to various stimuli, influencing isotype switching and differentiation into antibody-secreting cells (ASCs).
Purpose of the Study:
- To introduce a novel in vitro culture system for CTV-labeled human naïve B cells.
- To establish a method for precise, time-based data acquisition on B cell proliferation, division-linked isotype switching, and differentiation.
Main Methods:
- Utilized Cell Trace Violet (CTV) for cell division tracking.
- Developed a novel in vitro culture setup for human naïve B cells.
- Collected time-based data on proliferation and differentiation markers.
Main Results:
- The novel culture setup enables precise quantification of B cell proliferation and division kinetics.
- Distinct B cell responses to T cell-independent (CpG, anti-Ig) and T cell-dependent (CD40L, IL-21) stimuli were analyzed.
- Quantitative analysis revealed biologically meaningful measurements of division rounds and cell fate.
Conclusions:
- The described method provides valuable insights into the intrinsic programming of B cell health.
- This approach is applicable for clinical investigations assessing B cell responses.
- Precise time-based data on B cell proliferation, isotype switching, and differentiation can be obtained.
Abstract:
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.
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