AbstractSeveral anti-cancer therapies target the epidermal growth factor receptor (EGFR). Radionuclide imaging of EGFR expression in tumours may aid in selection of optimal cancer therapy. The ¹¹¹In-labelled DOTA-conjugated Z_EGFR:2377 Affibody molecule was successfully used for imaging of EGFR-expressing xenografts in mice. An optimal combination of radionuclide, chelator and targeting protein may further improve the contrast of radionuclide imaging. The aim of this study was to evaluate the targeting properties of radiocobalt-labelled DOTA-Z_EGFR:2377. DOTA-Z_EGFR:2377 was labelled with ⁵⁷Co (T_1/2 = 271.8 d), ⁵⁵Co (T_1/2 = 17.5 h), and, for comparison, with the positron-emitting radionuclide ⁶⁸Ga (T_1/2 = 67.6 min) with preserved specificity of binding to EGFR-expressing A431 cells. The long-lived cobalt radioisotope ⁵⁷Co was used in animal studies. Both ⁵⁷Co-DOTA-Z_EGFR:2377 and ⁶⁸Ga-DOTA-Z_EGFR:2377 demonstrated EGFR-specific accumulation in A431 xenografts and EGFR-expressing tissues in mice. Tumour-to-organ ratios for the radiocobalt-labelled DOTA-Z_EGFR:2377 were significantly higher than for the gallium-labelled counterpart already at 3 h after injection. Importantly, ⁵⁷Co-DOTA-Z_EGFR:2377 demonstrated a tumour-to-liver ratio of 3, which is 7-fold higher than the tumour-to-liver ratio for ⁶⁸Ga-DOTA-Z_EGFR:2377. The results of this study suggest that the positron-emitting cobalt isotope ⁵⁵Co would be an optimal label for DOTA-Z_EGFR:2377 and further development should concentrate on this radionuclide as a label.