Copy number variation (CNV) remains poorly defined in many populations, including Mexican Americans. We report the discovery and genetic confirmation of copy number variable regions (CNVRs) in subjects of the San Antonio Family Heart and the San Antonio Family Diabetes Gallbladder Studies, both comprised of multigenerational pedigrees of Mexican American descent. In a discovery group of 1677 participants genotyped using Illumina Infinium Beadchips, we identified 2937 unique CNVRs, some with observation frequencies as low as 0.002, using a process that integrates pedigree information with CNV calls made by PennCNV and/or QuantiSNP. Quantitative copy number values had statistically significant (P≤1.792e-5) heritability estimates ranging from 0.139 to 0.863 for 2776 CNVRs. Additionally, 920 CNVRs showed evidence of linkage to their genomic location, providing strong genetic confirmation. Linked CNVRs were enriched in a set of independently identified CNVRs from a second group of 380 samples, confirming that these CNVRs can be used as predefined CNVRs of high confidence. Interestingly, we identified 765 putatively novel variants that do not overlap with the Database of Genomic Variants. This study is the first to use linkage and heritability in multigenerational pedigrees as a confirmation approach for the discovery of CNVRs, and the largest study to date investigating copy number variation on a genome-wide scale in individuals of Mexican American descent. These results provide insight to the structural variation present in Mexican Americans and show the strength of multigenerational pedigrees to elucidate structural variation in the human genome.European Journal of Human Genetics advance online publication, 22 August 2012; doi:10.1038/ejhg.2012.188.