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Updated: Aug 8, 2026
The Use of Carboxyfluorescein Diacetate Succinimidyl Ester (CFSE) to Monitor Lymphocyte Proliferation
Published on: October 13, 2010
[Application of vital dye CFDA-SE and SNARF-1 to evaluate mixed lymphocyte reaction]
Jing-xian Zhao1, Yao-ying Zeng, Yi Liu
1Key Laboratory of Ministry of Education for Tissue Transplantation and Immunology, Jinan University, Guangzhou 510630, China. zhaojingxian@126.com
Insights
This study introduces a new flow cytometry method using CFDA-SE and SNARF-1 dyes for precise evaluation of responsive intensity in one-way mixed lymphocyte reactions (MLR). This technique enhances the analysis of immune cell proliferation and T-cell responses.
Area of Science:
- Immunology
- Cell Biology
Context:
- Mixed lymphocyte reaction (MLR) is crucial for studying T-cell mediated immune responses.
- Accurate quantification of T-cell proliferation in MLR is essential for understanding immune activation.
Purpose:
- To develop and validate a novel, comprehensive method for assessing responsive intensity in one-way mixed lymphocyte reactions (MLR).
Summary:
- Responder lymphocytes (CFDA-SE labeled) and stimulator splenocytes (SNARF-1 labeled) were used in a 96-hour mixed lymphocyte culture (MLC).
- Flow cytometry analysis, with specific gating strategies to exclude stimulator cells, enabled precise measurement of responder cell proliferation.
- ModFit software provided key proliferation indices, including precursor frequency and proliferation index.
Impact:
- This optimized flow cytometry approach offers a powerful and accurate tool for evaluating immune cell responsiveness in MLR.
- The method facilitates deeper insights into T-cell activation and immune dynamics, crucial for immunological research and diagnostics.
Aim:
To establish a new method for more comprehensive evaluation of responsive intensity in one-way mixed lymphocyte reaction (MLR).
Methods:
CFDA-SE labeled lymphocytes from lymph nodes of BALB/cJ mice were applied as responder cells, and SNARF-1 labeled splenocytes from BALB/cJ or C57BL/6 mice which had been pretreated with mitomycin C were applied as stimulator cells to establish mixed lymphocyte culture (MLC). After 96 h of culture, cells were harvested and analyzed by flow cytometry to acquire the proliferative information of SNARF-1 negative and CFSE positive staining responder cells. ModFit software was then used to get more proliferation related index.
Results:
With increasing division cycles, the CFSE fluorescence was diluted to the extent that the responder cells could not be discriminated from stimulator cells, which was resolved by gating SNARF-1 positive cells out to define responder cells. Several important proliferation related indexes were obtained by ModFit software, such as precursor frequency and proliferation index, etc.
Conclusion:
Our results show that application of vital dye CFDA-SE and SNARF-1, combined with flow cytometry is a powerful tool for evaluation of responsive intensity in one-way MLR.
