The assay was further applied in 13 ER+ breast cancer (BrCa) primary tumour tissues (FFPEs), 13 non-cancerous breast tissues (mammoplasties), and 32 pairs of liquid biopsy samples [circulating tumour cells (CTCs) and paired plasma circulating tumour DNA (ctDNA)] obtained at different time points from 8 ER+ metastatic breast cancer patients. Here, ESR1 is linked to breast cancer.