Immune activation may play an important role in the pathogenesis of acute rheumatic fever (ARF). The objective of the present study was to investigate serum concentrations of various markers of immune activation in ARF patients. Sera of 32 patients with ARF were investigated, 20 of them in follow-up. Radioimmunoassay was used to quantify neopterin and ELISA for the measurement of 55-kDa-type soluble tumor necrosis factor receptor (sTNF-R) and soluble interleukin-2 receptor (sIL2-R). Markers of immune activation were found to be raised in 48% (sTNF-R), 28% (sIL2-R), and 78% (neopterin) of patients at the onset of ARF. There were significant correlations between the concentrations of neopterin and sTNF-R (rs = 0.60, P < 0.001) or sIL2-R (rs = 0.35, P < 0.05). Higher neopterin concentrations were found in patients with combined aortic and mitral insufficiency than in patients with mitral valve lesions alone (U = 2.67, P < 0.05) or without valve lesion (U = 2.36, P < 0.05). Increased concentrations of neopterin, sTNF-R, and sIL2-R demonstrate activation of the cellular immune system in patients with ARF. Higher serum neopterin concentrations are associated with development of combined aortic and mitral insufficiency during the first episode of ARF.