Detection of H2O2 inhibition and genetic mutations using ChIn assays. (a) Reactive oxygen species signaling is critical for copper-induced inflammation. Diphenyleneiodonium (DPI), an inhibitor of NADPH oxidases, impairs wounds to leukocyte signaling, resulting in reduced leukocyte numbers at damaged neuromasts. Larvae at age 56 hpf were treated with 100 μM DPI for 1 hour prior to copper treatment (10 μM CuSO4 for 40 minutes). DPI treatment results in significantly lower leukocyte numbers within the myoseptum compared to untreated control larvae. (b) The ChIn assay enables detection of genetic mutations affecting an inflammatory response. Clutches from matings between homozygous Wiskott-Aldrich syndrome (was) gene males and heterozygous was females were treated with 10 μM CuSO4, fixed and scored by Sudan Black staining, and individual larvae were subsequently genotyped. Both homozygous and heterozygous was mutants recruited significantly lower numbers of leukocytes to the myoseptum than wild-type larvae upon copper-induced neuromast damage. In addition, the inflammatory response of homozygous mutants is significantly lower than that of heterozygous mutants. **P = 0.0045. ***P < 0.001.
d'Alençon et al. BMC Biology 2010 8:151 doi:10.1186/1741-7007-8-151