Alterations in DNA methylation have implicated as an epigenetic event in the pathogenesis of late-onset Alzheimer’s disease (LOAD). The objective of this work was to evaluate global DNA methylation levels for long interspersed nuclear element 1 (LINE-1) repetitive sequences in Colombian patients with LOAD and controls. The LINE-1 DNA methylation levels in peripheral blood samples from 28 Colombian patients with LOAD and 30 healthy participants were assessed using a methylation-sensitive high-resolution melting (MS-HRM) quantitative assay. We did not find differences in LINE-1 methylation levels between patients with Alzheimer’s disease (AD; median 76.2%, interquartile range [IQR]: 69.8-81.9) and control participants (median 79.8%, IQR: 73.2-83.8; P = .3). Additional stratified analyses did not show differences in LINE-1 methylation levels for male or female patients versus controls nor for apolipoprotein E4 carriers and noncarriers. This is the first report of LINE-1 methylation levels in patients with LOAD using the cost-effective MS-HRM technique, and this is the first global DNA methylation study in Latin American patients with AD.