One hundred sixty-one DRB1*03 positive individuals from each of five U.S. population groups (Caucasoids, African Americans, Asians/Pacific Islanders, Hispanics, and Native Americans) were randomly selected from a database of 82,979 individuals. DRB1*03 alleles were identified by polymerase chain reaction-sequence-specific oligonucleotide probe typing. A total of six DRB1*03 alleles out of 21 known alleles were detected. DRB1*03011 was the predominant DRB1*03 allele in all populations. Caucasoids were found to be the least diversified; only DRB1*03011 was observed. African Americans carried DRB1*03021 at a high frequency. This allele was observed in three other populations. DRB1*0304 was found in Asians/Pacific Islanders and DRB1*0305, DRB1*0307 and a new allele, DRB1*0316, was found in Hispanics. A subset of individuals was also typed for DRB3 alleles. DRB3*0101, DRB3*0202, and DRB3*0301 were detected and seven DRB1-DRB3 haplotypes were defined. Testing of other individuals not included in the DRB1*03 frequency study identified a variation of a common extended haplotype, A1, B8, DR3, which carries DRB1*0304 and two previously unreported DRB1*03 alleles, DRB1*0311 and *0320, are also described.