Abstract:

BACKGROUND: There have been no standard procedures regarding how to simply and efficiently separate and purify human peripheral blood monocytes and stimulate them into dendritic cells.
OBJECTIVE: To investigate the efficacy of purification of human peripheral blood monocytes on gelatin-coated surfaces, analyze the phenotype of dendritic cells generated by these monocytes, and make a comparison with conventional plastic adhesion method.
METHODS: Human peripheral blood mononuclear cells were harvested by Ficoll-Hypaque gradient centrifugation. Gelatin group and common plastic group were designated according to coating flasks with or without gelatin. Monocytes were harvested from each group and then were stimulated into dendritic cells. The number of monoctyes, CD14 positive rate of monocytes, contaminated T, B lymphocytes, the expression of CD1a, CD83 on immature and mature dendritic cells were determined. Cell viability was determined by trypan blue staining. Platelet contamination was compared between gelatin and common plastic groups.
RESULTS AND CONCLUSION: The number of monocytes and CD14 positive rate were significantly higher in the gelatin group than in the common plastic group (P < 0.05). Contaminated T, B lymphocytes were significantly more in the common plastic group than in the gelatin group (P < 0.05). There were no significant differences in cell viability and dendritic cell phenotype between gelatin group and common plastic group (P > 0.05). Platelet contamination rate was significantly lower in the gelatin group than in the common plastic group. Gelatin-coated surfaces can effectively and simply isolate monocytes and successfully stimulate them into mature dendritic cells.