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We have developed a high-throughput, multiplex reverse transcription PCR (RT-PCR) assay that is suitable for the analysis of medium- to low-copy cellular RNA transcripts from small numbers of cells (104). High throughput was attained by utilizing microplate-based RNA extraction and RT-PCR protocols, followed by PCR product visualization of a multiwelled agarose gel, stained with SYBR® Green I dye. The transcriptional assay was unaffected by solvents (dimethyl sulfoxide and methanol) routinely used in high-throughput drug screens at concentrations required for compound solubilization. Furthermore, it has been used successfully for the investigation of differential mRNA expression levels of tumor necrosis factor alpha (TNF-α) and Interleukin-1 beta (IL-1β) in lipopolysaccharide (LPS)stimulated THP-1 cells (a human monocytic cell line) and the identification of specific IL-1β transcriptional inhibitors.