Overexpression of vascular endothelial growth factor (VEGF) and VEGF receptors (VEGFRs) indicates poor prognosis for cancer patients in a variety of clinical studies. Our goal is to develop a tracer for positron emission tomography (PET) imaging of VEGFR expression using recombinant human VEGF121 with three lysine residues fused to the N-terminus (denoted as K3-VEGF121)， which can facilitate radiolabeling without affecting its VEGFR binding affinity. K3-VEGF121 was conjugated with 1，4，7-triazacyclononane-1，4，7-triacetic acid (NOTA) and labeled with (61)Cu (t1/2: 3.3 h; 62% β(+)). The IC50 value of NOTA-K3-VEGF121 for VEGFR-2 was comparable to that of K3-VEGF121 (1.50 and 0.65 nM， respectively) based on a cell binding assay. (61)Cu labeling was achieved with good yield (55 ± 10%) and specific activity (4.2 GBq/mg). Serial PET imaging showed that the 4T1 tumor uptake of (61)Cu-NOTA-K3-VEGF121 was 3.4 ± 0.5， 4.9 ± 1.0， 5.2 ± 1.0， and 4.8 ± 0.8%ID/g (n = 4) at 0.5， 2， 4， and 8 h postinjection， respectively， which was consistent with biodistribution data measured by γ counting. Blocking experiments and ex vivo histology confirmed the VEGFR specificity of (61)Cu-NOTA-K3-VEGF121. Extrapolated human dosimetry calculation showed that liver was the organ with the highest radiation dose. The use of (61)Cu as the radiolabel is desirable for small proteins such as K3-VEGF121， which has a much higher β(+) branching ratio than the commonly used (64)Cu (62% vs 17%)， thereby offering stronger signal intensity and lower tracer dose for PET imaging.