RT-PCR detection of IL-6 mRNA. RT-PCR was performed (35 cycles) to detect mRNA for β-actin and IL-6 from cells recovered from dual-chambered plates at 96 hours in which IL-6 protein levels were known. Primers were designed to cross intron/exon boundaries. Negative controls included omitting reverse transcriptase (not shown) from the reverse transcription reaction or using H₂O. Results from two independent experiments are presented. mRNA from osteoblasts only (HOB), mRNA from osteoblasts and CD34⁺ cells in coculture (contact), mRNA from osteoblasts recovered from dual-chambered coculture (HOB noncontact), and mRNA from CD34⁺ cells recovered from dual-chambered coculture (CD34⁺ non-contact).