Alpha 2-HS glycoprotein (AHSG) is a human plasma glycoprotein that exhibits genetic polymorphism on isoelectric focusing (IEF). To identify the origin of two common alleles, AHSG*1 and *2, we examined nucleotide exchanges in the gene. AHSG cDNA was obtained by RT-PCR from poly(A) RNA of seven liver tissue samples and subcloned into a plasmid vector. After sequencing, we found six single nucleotide differences in comparison with the originally reported sequence. In particular, the nucleotide substitutions of C to T at amino acid position 230 and C to G at position 238 were common among the samples exhibiting phenotype 2-1 or 2. Since these substitutions might give rise to a NlaIII site and a SacI site, respectively, for the potential AHSG*2, we analyzed these substitutions by PCR-RFLP using genomic DNA of 68 individuals. The result was consistent with the IEF analysis of the corresponding serum, indicating that AHSG*1 was characterized by ACG (Thr) at position 230 in exon 6 and ACC (Thr) at position 238 in exon 7, and that AHSG*2 was characterized by ATG (Met) at position 230 and AGC (Ser) at position 238.