Supplementary Material for: Toll-Like Receptor Responses in IRAK-4-Deficient Neutrophils van Bruggen R. A.Drewniak A.T.J.Tool M.Jansen van Houdt M. J.Geissler van den Berg T.K. H.Chapel T.W.Kuijpers 2009 Human neutrophils were found to express all known Toll-like receptors (TLRs) except TLR3 and TLR7. IRAK-4-deficient neutrophils were tested for their responsiveness to various TLR ligands. Essentially all TLR responses in neutrophils, including the induction of reactive oxygen species generation, adhesion, chemotaxis and IL-8 secretion, were found to be dependent on IRAK-4. Surprisingly, the reactivity towards certain established TLR ligands, imiquimod and ODN-CpG, was unaffected by IRAK-4 deficiency, demonstrating their activity is independent of TLR. TLR-4-dependent signaling in neutrophils was totally dependent on IRAK-4 without any major TRIF-mediated contribution. We did not observe any defects in killing capacity of IRAK-4-deficient neutrophils for <i>Staphylococcus aureus</i>, <i>Escherichia coli</i> and <i>Candida albicans</i>, suggesting that microbial killing is primarily TLR independent.