TF and ovarian neoplasm: We have characterized several receptor‐probe interactions, such as transferrin‐transferrin receptor,[14, 16, 20, 33, 34] Cetuximab‐Epidermal Growth Factor Receptor,[22] and TZM‐HER2,[21, 35] providing an accurate measurement of bound and unbound fraction of the probe to its respective target.[16, 21, 22] Herein, we present dark quencher‐based MFLI FRET as a unique noninvasive approach to measure the TZM‐HER2 drug‐target engagement of HER2‐positive human breast and ovarian tumor xenografts mice model, which display varying TMEs.