To test whether the presence of indolyl-3-acryloylglycine (IAG) is associated with autism, we analyzed urine from population-based, blinded cohorts. All children in York, UK with autism spectrum disorders (ASDs), diagnosed using ICD-10 research diagnostic criteria, were invited to participate. Fifty-six children on the autism spectrum (mean age 9y 8mo, SD3y 8mo; 79% male) agreed to participate, as did 155 children without ASDs (mean age 10y, SD3y 2mo; 54% male) in mainstream and special schools (56 of whom were age-, sex-, and school-matched to children with ASDs). IAG was found at similar levels in the urine of all children, whether IAG concentrations or IAG:creatinine ratios were compared. There was no significant difference between the ASD and the comparison group, and no difference between children at mainstream schools and those at special schools. There is no association between presence of IAG in urine and autism; therefore, it is unlikely to be of help either diagnostically or as a basis for recommending therapeutic intervention with dietary manipulation. The significance of the presence of IAG in urine has yet to be determined.