RNA sequence requirements for immunostimulation by protamine-RNA particles. Large nanoparticles were produced using different RNA oligonucleotides at 1 mg/mL in water, and Protamine 1000 diluted 2.5 times in water. Resulting particles were approximately 500 nm in diameter. Fresh human PBMCs were added and cultured for 18 hours. Supernatants were then tested for TNF-α and interferon-α content by ELISA. (A) The effect of the RNA length on immunostimulation was tested. (B) The effect of the RNA sequence on immunostimulation was tested. Each set of experiments was repeated 3 times. Representative data are shown.