Human cytochrome P450 3A4 (CYP34A) plays an important role in the metabolism of many endo- and xenomaterials. It also exhibits a substantial interindividual variation in enzymatic activity. It has been shown that the mutant alleles of CYP3A4 encoding inactive/decreased enzymes are largely caused by single nucleotide polymorphisms (SNPs) in the gene sequence. In the present study, with the goal of detecting the known SNPs of CYP3A4, an oligonucleotide microarray was created. A genotyping standard for this microarray was also established using constructed plasmids as standard templates. The 12 SNPs of CYP3A4 in 387 Chinese DNA samples were screened using this oligonucleotide microarray. Three heterozygous subjects of CYP3A4*/*4, 5 heterozygous subjects of CYP3A4*1/*5, 4 heterozygous subjects of CPY3A4*1/6, and 6 heterozygous subjects of CYP3A4*1/*18 were found. The genotyping results of the 18 heterozygous subjects and 12 wild-type subjects were validated by direct sequencing.