[In vitro cultivation of dendritic cells with serum-free medium].
This study was aimed to investigate the protocol in vitro to incubate the dendritic cell (DC) derived from peripheral blood monocytes using serum-free medium X-VIVO 20. Peripheral blood monocytes from healthy donors were treated with 100 ng/ml GM-CSF and 500 U/ml IL-4, respectively. After cultivation for 6 days, they were treated with 100 ng/ml calcium ionophore A23187. After cultivation for 24 hours the cellular morphology was observed under invert microscope, the surface markers were analyzed by flow cytometry, the proliferation of allogenetic T cells was detected by MTT colorimetry, the specific cytotoxicity of T cells primed with DC was examined by MTT assay. The results showed that in all three groups with serum-free, fetal calf serum (FCS) and human AB serum mediums, cells displayed characteristic morphological features of DC. Simultaneously CD14 expression was decreased, and CD83, HLA-DR and CDw123 expression were increased on these cells. In addition, DCs cultured with these methods could evidently stimulate the proliferation of allogenetic T cell. As compared with the two controls of serum containing groups, the cultured cells in the serum-free groups showed almost the same allo-stimulatory capability and cellular morphology and surface markers, and T lymphocytes primed with the culture-derived DC exhibited the similar killing activity to K562 (P > 0.05). It is concluded that there is no significance in DC numbers, morphology, epitope and ability to stimulate the proliferation of allogenetic T cells between DC induced by serum-free X-VIVO 20 medium and DC induced by serum-contained medium. DC cultured and induced by serum-free medium is worth using in practice widely.
Zhang
Huai-Dong
HD
Department of Hematology, Shanton Central Hospital, Shanton 310003, China. zhd1977@163.com
Song
Zhen-Lan
ZL
Li
Wei-Ping
WP
D018929:Cell Culture Techniques; D016895:Culture Media, Serum-Free; D003713:Dendritic Cells / Q000166:cytology*; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology*; D006801:Humans; D015847:Interleukin-4 / Q000494:pharmacology; D009000:Monocytes / Q000166:cytology*; D011994:Recombinant Proteins; D013601:T-Lymphocytes / Q000166:cytology; D013602:T-Lymphocytes, Cytotoxic
D016895:Culture Media, Serum-Free; D011994:Recombinant Proteins; D015847:Interleukin-4; D016178:Granulocyte-Macrophage Colony-Stimulating Factor