1. Analysis of TNF-α gene transcription and mRNA accumulation in THP-1 cells.
Analysis of TNF-α gene transcription and mRNA accumulation in THP-1 cells. (A) Nuclear run-on assay for newly transcribed TNF-α mRNA. Differentiation into macrophages was induced by incubation of the cells with 200 nM PMA for 24 h. Nuclei were harvested from LPS-stimulated and nonstimulated cells at different intervals, and in vitro transcription of the TNF-α gene was analyzed with specific probes for TNF-α. β-Actin was used as the internal control, and pGEM 7 vector was used as the negative control. (B) Densitometric analysis of the bands in panel A. (C) Kinetics of TNF-α mRNA accumulation in LPS-stimulated differentiated and undifferentiated THP-1 cells. Cells were stimulated with 100 ng of LPS per ml, and mRNA was extracted in the indicated time intervals. TNF-α mRNA was quantified by RT-PCR with [32P]dCTP in relation to a standard curve.
Shogo Takashiba et al. Infect. Immun. 1999; doi: