PURPOSE: The purpose of this study was to investigate the albumin-binding compound 111In-C4-DTPA as an imaging agent for the detection of endogenous albumin accumulation in tumors. METHODS: 111In-C4-DTPA was injected in healthy nude mice for pharmacokinetic and biodistribution studies (10 min, 1, 6, 24, and 48 h, n = 4) and subsequently in tumor-bearing mice for single-photon emission computed tomography/X-ray-computed tomography (SPECT/CT) imaging studies. Four different human tumor xenograft models (LXFL529, OVXF899, MAXFTN401, and CXF2081) were implanted subcutaneously unilaterally or bilaterally (n = 4–8). After intravenous administration of 111In-C4-DTPA, SPECT/CT images were collected over 72 h at 4–6 time points. Additionally, gamma counting was performed for the blood, plasma, lungs, heart, liver, spleen, kidneys, muscle, and tumors at 72 h post-injection. RESULTS: 111In-C4-DTPA bound rapidly to circulating albumin upon injection, and the radiolabeled albumin conjugate thus formed was stable in murine and human serum. SPECT/CT images demonstrated a time-dependent uptake with a maximum of 2.7–3.8% ID/cm3 in the tumors at approximately 24 h post-injection and mean tumor/muscle ratios in the range of 3.2–6.2 between 24 and 72 h post-injection. The kidneys and bladder were the predominant elimination organs. Gamma counting at 72 h post-injection showed 1.3–2.5% ID/g in the tumors and mean tumor/muscle ratios in the range of 4.9–9.4. CONCLUSION 111In-C4-DTPA bound rapidly to circulating albumin upon injection and showed time-dependent uptake in the tumors demonstrating a potential for clinical application as a companion imaging diagnostic for albumin-binding anticancer drugs.