Atopic dermatitis is a disease with an impaired skin barrier that affects 15%–20% of children. In the normal epidermis, the stratum corneum chymotryptic enzyme (SCCE) thought to play a central role in desquamation by cleaving proteins of the stratum corneum (e.g., corneodesmosin and plakoglobin). Genetic variations within the SCCE gene could be associated with dysregulation of SCCE activity leading to an abnormal skin barrier. We screened the SCCE gene for variations and performed a case–control study on 103 atopic dermatitis patients and 261 matched controls. 16 synonymous single nucleotide polymorphisms (SNPs) have been identified and a 4 bp (AACC) insertion has been found in the 3′UTR. We performed an association study of the SCCE AACC insertion in the 3′UTR, and found a significant trend between the AACC allele with the two insertions and disease in the overall data set [odds ratio (OR)=2.31; p=0.0007]. The AACC insertion in the SCCE gene may result in a change to SCCE activity within the skin barrier. These findings suggest that SCCE could have an important role in the development of atopic dermatitis.