To assess PSMA targeting, PSMA-expressing cells DU145-PSMA and non-PSMA-expressing cells DU145 cells were seeded in 6-well plates at a density of 0.5 × 10 cells/well 1 day prior to the experiment. Three technical replicates were performed for each condition. Cell medium was replaced with 1 mL complete medium 1 hour before the cells were treated. [ 201 Tl(K222-PSMA)] + (50 kBq, in 10 uL of phosphate buffered saline) was added to each well, and the cells incubated at 37 ˚C for 15 min and 1 h. Competition studies were also performed following coincubation with the PSMA-inhibitor 2-(phosphonomethyl)pentane-1,5-dioic acid (PMPA; 30 μL of 750 mM PMPA solution/well). After incubation, plates were placed on ice, the supernatant was removed and the cells were washed with ice cold phosphate buffered saline solution (2 × 1 mL). The cells were lysed with ice cold radioimmunoprecipitation assay (RIPA) buffer (Thermo Fisher Scientific Inc., 500 μL), and radioactivity content was determined by gamma counter (1282 Compugamma; LKB, window set to channels 35-110 to measure 201 Tl gamma emissions). Cell uptake was also performed following [ 201 Tl]TlCl incubation (50 kBq, 10 μL) for 15 min and 1 h. Cells were harvested and radioactivity measured as described above.