Imaging of β-amyloid (Aβ) plaques in the brain may facilitate the diagnosis of cerebral β-amyloidosis, risk prediction of Alzheimer's disease (AD), and effectiveness of anti-amyloid therapies. The purpose of this study was to evaluate novel (123)I-labeled pyridyl benzofuran derivatives as SPECT probes for Aβ imaging. The formation of a pyridyl benzofuran backbone was accomplished by Suzuki coupling. [(123)I/(125)I]-labeled pyridyl benzofuran derivatives were readily prepared by an iododestannylation reaction. In vitro Aβ binding assays were carried out using Aβ(1-42) aggregates and postmortem human brain sections. Biodistribution experiments were conducted in normal mice at 2, 10, 30, and 60 min postinjection. Aβ labeling in vivo was evaluated by small-animal SPECT/CT in Tg2576 transgenic mice injected with [(123)I]8. Ex vivo autoradiography of the brain sections was performed after SPECT/CT. Iodinated pyridyl benzofuran derivatives showed excellent affinity for Aβ(1-42) aggregates (2.4 to 10.3 nM) and intensely labeled Aβ plaques in autoradiographs of postmortem AD brain sections. In biodistribution experiments using normal mice, all these derivatives displayed high initial uptake (4.03-5.49% ID/g at 10 min). [(125)I]8 displayed the quickest clearance from the brain (1.30% ID/g at 60 min). SPECT/CT with [(123)I]8 revealed higher uptake of radioactivity in the Tg2576 mouse brain than the wild-type mouse brain. Ex vivo autoradiography showed in vivo binding of [(123)I]8 to Aβ plaques in the Tg2576 mouse brain. These combined results warrant further investigation of [(123)I]8 as a SPECT imaging agent for visualizing Aβ plaques in the AD brain.