Beyond its linear sequence, DNA folds into higher-order structures that add a dynamic layer of regulatory control within the genome. Among these, G-quadruplexes (G4s) and i-motifs (iMs) are non-canonical DNA structures known to influence gene expression. G4s and iMs form within guanine- and cytosine-rich regions, respectively, and are often located near oncogenes and regulatory elements, suggesting their involvement in cancer progression. Although previous research has demonstrated the interdependent nature of G4 and iM formation, no studies have directly visualized both structures within the same cells. Here, we present the first co-staining and co-visualization of G4s and iMs within the same cells to investigate their dynamics during cell cycle progression. Using G4-specific and iM-specific antibodies, we assess colocalization and formation patterns in different breast cancer subtypes. Beyond conventional confocal microscopy, expansion microscopy enables high-resolution visualization of their spatial distribution within nuclei. To further probe their relationship, drug perturbation experiments using small-molecule ligands that selectively stabilize G4s or iMs can be performed. This will allow us to determine how structural stabilization influences their colocalization patterns. Ultimately, this study will provide fundamental insights into the dynamics and colocalization of these structures within breast cancer cells.