Background: The relationship of juvenile ankylosing spondylitis(JAS) and juvenile rheumatoid arthritis(JRA) with HLA-B27 alleles has been studied for about 25 years. Some researchers abroad reported that the relationship between ankylosing spondylitis and HLA-B27 allele subtypes is different from that between arthritis and HLA-B27 allele subtypes. Objective: To investigate the relationship of JAS and JRA with HLA-B27 allele subtypes and its role in the pathogenesis so as to find out the testing methods for earlier diagnosis of JAS or JRA. Design: Non-random control laboratory study. Setting, participants and interventions: A total of 60 patients diagnosed with JAS or JRA in Beijing Children's Hospital were recruited, including 5 fathers or mothers from 5 patients' families, compared with 9 normal controls (from 200 donors for bone marrow transplantation). We studied 74 cases of HLA-B27 allele subtypes and analyzed their correlation by using PCR/SSP. Main outcome measures: Correlation of JAS and JRA with HLA-B27 allele subtypes; difference of HLA-B*2704 between JAS and JRA; data of patients' families. Haplotypes analysis and homozygote analysis on patients' families showed that there existed a close link between HLA-B* 2704 and C*1202, but not with HLA-A, and that JAS had a remarkable tendency of family inheritance. Results: HLA-B*2704, *2705, *2702 and *2707 were found in the cases. In JAS group, allelic frequency of HLA-B27 was B*2704 56. 25%, B*2705 40.63% and B*2702 3.13% , respectively. However, in JRA group, it was B*2705 60.7%, B* 2704 28.57%, B*2702 3.57% and B* 2707 7.14%, respectively. Comparison between JAS and JRA showed that the allelic frequency of HLA-B*2704 in JAS patients was higher than that in JAS patients (RR = 3.21, P < 0.05). Conclusion: The susceptibility of JAS is positively associated with subtypes HLA-B*2704. Testing HLA-B27 subtypes may be a valuable marker in differential diagnosis of AS and JRA.