Title: Application of the proliferative response of human T lymphocyte subsets labeled by CFSE in virus infectious diseases
Abstract: Objective To study the methodology of combining carboxyfluorescein diacetatesuccinimidyl ester (CFSE) with flow cytometry to label the proliferitive response of human T lymphocyte subsets and its application in virus infectious diseases. Methods To select the optimal concentration and culture time, the fresh PBMC isolated from healthy subjects were labeled with CFSE on different concentrations, divided into unstimulated group and stimulated group, one group added with CD28, the other group added with CD28 and PHA, and incubated at different time from 5 to 7 days, then stained and determined by flow cytometry.The proliferative dynamics of human T cell subsets were analyzed with CellQuest and ModFit software.At the above optimal experiment conditions, the proliferative situation of T lymphocyte subsets from CMV-IgG positive and HLA-A2 positive subjects after simulated with CMV-pp65 peptide were analyzed. Results The optimal labeling concentration was 0.5 μmol/L for CFSE and 2 μg/ml for PHA.Under these conditions, six day was selected as the optimal culture time.The proliferation of T lymphocyte subsets were atypically asynchronous after stimulated with PHA, and the CD8+T cells showed obvious proliferation after simulated with CMV-pp65 peptide.The combination of CellQuest with ModFit software can realize the quantification and visualization in analyzing the proliferation of lymphocyte. Conclusion CFSE staining combined with fluorescent antibody labeling and cytometry are powerful tools for analysis of lymphocytic Proliferation.It can effectively detect the proliferation kinetics of T lymphocyte subsets after stimulated with virus antigen.It will be of great value in exploring the pathogenesis of virus infectious disease.
Publication Year: 2008
Publication Date: 2008-01-01
Language: en
Type: article
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