ABSTRACT Background and Purpose This study aims to investigate the longitudinal changes in translocator protein (TSPO) following stroke in different brain regions and potential associations with chronic brain infarction. Methods Twelve patients underwent SPECT using the TSPO tracer 6‐Chloro‐2‐(4ʹ‐123I‐Iodophenyl)‐3‐( N , N ‐Diethyl)‐Imidazo[1,2‐a]Pyridine‐3‐Acetamide, as well as structural MRI, at 10, 41, and 128 days (median) after ischemic infarction in the middle cerebral artery. TSPO expression was measured in lesional (MRI lesion and SPECT lesion), connected (pons and ipsilesional thalamus), and nonconnected (ipsilesional cerebellum and contralesional occipital cortex) regions. Correlations were explored between the volume of chronic infarction and TSPO expression in nonconnected regions of interest (ROIs) at 128 days Results Throughout the study period, TSPO levels decreased by 24%–33% in lesional ROIs, while levels increased in connected ROIs by 35%–69% and in nonconnected ROIs by 53%–77%. At 128 days poststroke, TSPO expression in ipsilesional cerebellum positively correlated with chronic infarction volume ( p = 0.002, r 2 = 0.72). Conclusions This study expands the current knowledge of spatial and temporal TSPO expression in humans by quantifying TSPO changes in lesional, connected, and nonconnected brain regions at three time points after cerebral infarction as well as correlating late‐stage TSPO upregulation and chronic infarction volume.