Protein expression data suggests that the SPANX gene family is expressed throughout spermatogenesis, including post-meiotic expression, consistent with a potential role in sperm development. The genomic architecture of this region is unstable and past studies have found evidence of variation in this gene family. This study used a novel assay to evaluate copy number variation (CNV) in SPANX gene family, in fertile and infertile men. The case group was comprised of 50 oligozoospermic and 50 azoospermic men, and the control group was comprised of 67 normozoospermic men with children. The assay, real-time quantitative PCR, evaluated CNV of the entire gene cluster containing all four SPANXA-E genes and with SPANXB, found exclusively in maturing sperm. While variation was found in both groups, average CNV patterns did not differ between fertile and infertile males. As this was a targeted assay, it was limited in scope to detect further CNV at a genome-wide level which is an area of increasing interest in the field of genomics.