Bovine leukocyte adhesion deficiency (BLAD) is an autosomal recessive disease caused by A→G mutation in the CD18 gene. The disease is characterized by greatly reduced expression of the heterodimeric β2 integrin adhesion molecules on leukocytes, resulting in multiple defects in leukocyte function and significant deficits on performance traits in Holstein cattle. In this study, primers were designed to amplify the CD18 gene fragment and BLAD was detected by PCR-SSCP in a simple, highly accurate and high throughput manner. Results were confirmed by DNA sequencing. This study provides a more reliable and useful method for extensive screening of BLAD and also offers a theoretical basis for molecular diagnosis in Holstein calves.