Related Experiment Videos
In vitro cytokine mRNA expression in normal human peripheral blood mononuclear cells
S Barth1, B Kleinhappl, A Gutschi
1Institute of Hygiene, University of Graz, Austria. sonja.barth@kfunigraz.ac.at
Objective And Design:
Normal human peripheral blood mononuclear cells (PBMC) were analyzed for basic profiles of mRNA expression of distinct genes during incubation in a standard cell culture system.
Material:
Human PBMC from healthy adult blood donors.
Methods:
Steady-state mRNA expression was measured using a semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) assay.
Results:
Shortly after isolation and cell seeding, mRNA levels of monocyte-derived cytokines (IL-1alpha, IL-6, and TNF-alpha) were significantly increased, whereas lymphocyte-derived cytokines (IL-4, IFN-gamma) were not affected. Expression levels of monokines returned to basal within 24 h. At later stages of culture, the mRNA levels of all genes studied gradually increased and were significantly elevated after 96 h of incubation.
Conclusions:
Monokine and lymphokine mRNAs respond differently to cell culture even under control conditions. With regard to the enhanced mRNA expression of distinct cytokines in the early stages of culture, human PBMC should not be used for gene expression studies in vitro within 24 h of isolation.