Aptamer-integrated DNA tetrahedral sensors for cancer cell imaging and invasiveness assessment

DNA tetrahedron is a nanoscale framework self-assembled from DNA strands. Their nucleic acid composition confers excellent biocompatibility, rendering them highly suitable for drug delivery and biomarker imaging applications. Aptamers are short oligonucleotides that bind target proteins with high affinity and specificity. Integration of multiple aptamers into a DNA tetrahedron enhances sensor precision, facilitates targeted cell recognition, and promotes efficient cellular uptake. In this study, we developed a DNA tetrahedral aptasensor for selective imaging of cancer cells and assessment of their invasive potential (TDNA-IA). The sensor incorporates three aptamer domains targeting EpCAM, PTK7, and vimentin, respectively. The EpCAM- and PTK7-specific aptamers engage overexpressed surface markers on the membrane. The AND-logic fluorescence activation occurs exclusively in cells co-expressing both EpCAM and PTK7, thereby supporting high-specificity single-cell imaging. Experimental results demonstrate that the sensor effectively discriminates among 4T1, HeLa, and L929 cell lines based on fluorescence intensity, confirming its capability for specific imaging of cancer cells. Furthermore, the sensor features a hairpin-structured vimentin aptamer. The aptamer opens upon binding intracellular vimentin, restoring fluorescence and enabling quantitative analysis of vimentin expression. In vitro assays validated the sensor's high sensitivity, achieving a limit of detection (LOD) of 1.1 nM for vimentin. As a key epithelial-mesenchymal transition (EMT) biomarker, elevated vimentin expression signifies mesenchymal phenotype acquisition and heightened potential for invasion and metastasis. The developed sensor reliably quantifies intracellular vimentin levels, thereby providing an accurate assessment of cellular invasiveness.