Abstract
This research objective was to exploit a novel method for measuring the proliferation, cytotoxicity of cytokine-induced killer (CIK) cells using carboxyfluorescein succinimidyl ester/proliferation index (CFSE/PI) and flow cytometric assay. As cells divide, CFSE is apportioned equally between the two daughter cells, leading to a characteristic flow cytometric profile where a number of peaks of progressively halving CFSE fluorescence intensity were observed. The test principle of cytotoxicity is based on target cell labeling with CFSE and subsequent DNA-labeling with PI for the identification of target cells with compromised cell membranes. Results of CFSE fluorescence profile of the entire cell population showed that these cells underwent up to 6 divisions after 21 days. The percentage of total dividing cell population was 97.12%, of which 17.28% cells went through 6 rounds and 6.84% cells entered the seventh generation. The cytotoxicity of CIK cells showed a significant and positive correlation with effector: target (E:T) ratio ranging from 50:1 to 6.25:1. CFSE labeling combined with flow cytometry can therefore be used to monitor the proliferation of CIK cells. Moreover, it presents an advantageous method to measure cytotoxicity of CIK cells by avoiding radioactive substance, increasing sensitivity at low E:T ratio, analyzing on a cell-to-cell basis and allowing for long-term incubation. Key words: Flow cytometry, CFSE, cytokine-induced killer cell, proliferation, cytotoxicity.
Highlights
Cytokine-induced killer (CIK) cells are a group of heterogeneous cell populations derived from the human peripheral blood mononuclear cells (PBMCs) co-cultured in the presence of a variety of cytokines such as interleukin-2 (IL-2), interleukin-1 (IL-1α) and interferongamma (IFNγ), as well as monoclonal antibody against# Both authors contributed to the work.CD3
The profile position of carboxyfluorescein succinimidyl ester (CFSE) stained without induced control reduced in parent generation of cytokine-induced killer (CIK) cells. These results indicate that the division of CIK cells was monitored by CFSE labeling combined with flow cytometry
We explored the potential utility of an alternative nonradioisotopic marker of cell, the cytoplasmic dye carboxyfluorescein diacetate succinimidyl ester (CFDA-SE) combined with flow cytometry, to develop a novel method for tracking in real-time the number of divisions each CIK cell has undergone, and assessing accurately the phenotype and cytotoxicity of CIK cells
Summary
Cytokine-induced killer (CIK) cells are a group of heterogeneous cell populations derived from the human peripheral blood mononuclear cells (PBMCs) co-cultured in the presence of a variety of cytokines such as interleukin-2 (IL-2), interleukin-1 (IL-1α) and interferongamma (IFNγ), as well as monoclonal antibody against# Both authors contributed to the work.CD3 (anti-CD3mAb). Cytokine-induced killer (CIK) cells are a group of heterogeneous cell populations derived from the human peripheral blood mononuclear cells (PBMCs) co-cultured in the presence of a variety of cytokines such as interleukin-2 (IL-2), interleukin-1 (IL-1α) and interferongamma (IFNγ), as well as monoclonal antibody against. CIK cells are endowed with both a powerful anti-tumor T-lymphocyte activity and non-major histocompatibility complex-restricted in target cell recognition without any impact on bone marrow hematopoietic system (Schmidt-Wolf et al, 1997; Linn and Hui, 2003). CIK cells are a unique population of cytotoxic T lymphocytes (CTL) with the characteristic CD3+CD56+ phenotype (Alvarnas et al, 2001). The number of CD3+CD56+ lymphocytes is rare in human peripheral blood lymphocytes, these cells demonstrate dramatic
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