Abstract The oestrogen receptor (ER) is an important tumour target for molecular imaging and radionuclide therapy due to its overexpression in many malignant cells as compared to normal cells. Aiming to find new functional molecular imaging/therapeutic agents for ER positive tumours, we have synthesized a new estradiol derivative substituted at the 16-α position with a diethylene triamine tetraacetic acid (DTTA)-like chelating ligand through a four-carbon spacer. The new bioconjugate (H₄L), was used to synthesize the corresponding indium complexes (InL/[¹¹¹In]L). The radioactive complex [¹¹¹In]L was prepared in high yield (>98%) at final concentrations of 1 · 10 ^–4 M and its chemical identity was ascertained by comparing its HPLC gamma-chromatogram to the HPLC UV-vis-chromatogram of the InL analogue. [¹¹¹In]L is hydrophilic and kinetically stable in the presence of an excess of apo-transferrin and in human blood serum. Cellular studies in breast cancer cells (MCF-7 and MDA-MB-431) suggest that [¹¹¹In]L uptake may be mediated by an ER dependent mechanism. Biodistribution studies were performed in mice indicating a rapid clearance from most organs and a slow total excretion that occurs mainly by hepatobiliar pathway. High in vivo stability of [¹¹¹In]L was confirmed by HPLC analysis of urine and blood samples. Nevertheless, the hydrophilicity, the low ER affinity and the biodistribution of [¹¹¹In]L indicate that structural modifications are required to improve its behaviour for ER targeting in vivo.