RPA analysis of mRNA expression of cytokines and chemokines in monocytes derived from healthy and SCD patients. Peripheral blood monocytes were isolated from the whole blood of SCD patients and healthy individuals. RNA was isolated using TriZOL reagent, and 2 μg RNA from each sample was analyzed by RNase protection assay using a Riboquant Multi-Probe RNase protection assay system (Pharmingen). RPA was performed with riboprobes for (A) TNF-α, (B) IL-1β, (C) MIP-1β, (D) MCP-1, (E) IL-8, and (F) GAPDH.