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Updated: Aug 15, 2026
Optimized Protocol for Efficient Transfection of Dendritic Cells without Cell Maturation
Published on: July 8, 2011
[Immunostimulatory effect of human peripheral blood dendritic cells transfected by adeno-associated virus containing
Lei Xu1, Rong-cheng Luo, Chang-xuan You
1Department of Oncolagy, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China.
Objective:
To observe the immunostimulatory effect of human peripheral blood dendritic cells (DCs) transfected by Her2/neu gene delivered by adeno-associated virus.
Methods:
The mononuclear cells in healthy donors were isolated by Ficol-Hypaque density gradient separation and divided into transfection group and control group without transfection by the recombinant virus. The cells were initially cultured in RPMI 1640 medium supplemented with 10% AB human serum, followed by addition of granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin (IL)-4 and tumor necrosis factor (TNF)-alpha into the medium. The surface markers of DC were detected by flow cytometry, allogeneic mixed lymphocyte reaction assayed by 3H-TdR incorporation and the specific killing activity of T cells evaluated by MTT assay.
Results:
The expression rates of CD1a, CD86 and CD83 of the transfected DCs were 98.10%, 99.42%, and 84.59%, and those of the non-transfected DCs were 92.69%, 98.07%, and 82.72%, respectively, showing no obvious differences between the two groups. The rates of CD4 and CD80 expression were 61.02% and 97.61%, respectively, in the transfected DCs, significantly higher than those in the non-transfected cells (36.19% and 55.5%). Both groups of DCs stimulated a strong T cell proferation response. The transfected DCs were capable of inducing specific killing of the target tumor cells, with the highest killing rate of (39.7+/-7.2)%.
Conclusion:
The immunostimulatory effect of human peripheral blood DCs can be enhanced by Her2/neu gene transfection mediated by adeno-associated virus.
