The prevalence of intracranial arterial stenosis (IAS) as well as antinuclear antibody (ANA) positivity was found to be higher in Asians than that in the Western population. To investigate the relation of ANAs with IAS in patients with acute ischemic cerebrovascular disease, we enrolled 2492 patients with acute ischemic stroke or transient ischemic attack into the study. All the patients were categorized into 3 groups according to the IAS burden. Multinomial logistic regression analyses were used in statistical analysis. The positive rate of ANAs in the IAS ≥ 2 group was higher than that in the single IAS group and the no IAS group (p<0.001). The adjusted odds ratio (OR) for IAS ≥ 2 in ANAs-positive patients was 3.737 (95%CI=2.676-5.220, p<0.001) compared with the ANAs-negative patients. ANAs were associated with multiple IAS rather than single IAS in both male and female subgroups. Besides, ANAs were significantly associated with single and multiple IAS in individuals ≤ 60 years. However, ANAs were only associated with two or more IAS in two age groups (between 61 to 75 years and >75 years old). In summary, ANAs are associated with IAS in patients with acute ischemic cerebrovascular disease.