Mansuri, ShabnamShabnamMansuriOjha, SubhadraSubhadraOjhaKanvah, SriramSriramKanvah2025-08-312025-08-312025-03-1310.1002/adom.2024027582-s2.0-86000435239https://d8.irins.org/handle/IITG2025/28227Breast cancer remains the second most common cause of cancer-related deaths in women worldwide, with ≈70% of cases linked to the overexpression of Estrogen Receptor (ERα). Existing imaging tools often fail to reliably differentiate between ER-positive and ER-negative cancer cells. To address this limitation, two novel fluorescent probes, E2N and E2R, are synthesized by conjugating estradiol to styryl and rhodamine-based fluorophores using click chemistry. These probes are characterized by their photophysical properties, biocompatibility, and selective targeting of ER-positive cells. Cellular uptake studies demonstrate preferential internalization of E2N and E2R in ER-positive MCF-7, ZR-75-1, and T-47D cells, with minimal uptake in ER-negative MDA-MB-231, MDA-MB-468, and healthy COS-7 and NIH-3T3 cell lines. Kinetic studies reveal efficient and rapid uptake of E2N in ER-positive MCF-7 cells, while mechanistic investigations identified clathrin-mediated endocytosis as the receptor-mediated pathway for both probes. Localization studies further confirm their mitochondrial specificity in ER-positive cells, with E2R displaying higher mitochondrial selectivity. These findings underscore the potential of E2N and E2R as powerful tools for distinguishing ER-positive from ER-negative breast cancer cells. Their receptor-mediated targeting and precise imaging capabilities make them promising candidates for advancing breast cancer diagnostics and enabling more targeted therapeutic strategies.falseBreast cancer | Click chemistry | Estrogen Receptor (ERα) | Fluorescent probesArticle2195107113 March 202502402758arJournal0WOS:001434604500001