We sought to investigate specific hemostasis activation markers during electrophysiologic study (EPS) with consequent radiofrequency catheter ablation (RFA). Sixty patients were studied prospectively during routine EPS with RFA for paroxysmal supraventricular tachycardia. Blood samples were drawn before the insertion of venous sheaths (T0), at the end of EPS (T1), and 30 min after completion of RFA (T2). To study coagulation and fibrinolytic and platelet activity, we measured concentrations of thrombin-antithrombin III (TAT), D-dimers (DD), plasminogen activator inhibitor type 1 (PAI-1), tissue-type plasminogen activator (t-PA), and circulating platelet aggregates. The results are expressed as median and show 95% confidence levels. Levels of DD increased from 0.24 mg/L at T0 to 0.37 mg/L at T1 (P < 0.001) and to 0.59 mg/L at T2 (P < 0.001). TAT levels increased from 5.29 μg/L at T0 to 35.80 μg/L at T1 (P < 0.001) and decreased to 26.30 μg/L at T2 (P < 0.001). PAI-1 concentration decreased from 30.10 μg/L at T0 to 26.4 μg/L at T1 (P < 0.001). t-PA at T2 increased to 5.10 μg/L from 4.75 μg/L at T1 (P = 0.001). No other differences between corresponding medians were statistically significant (P > 0.05). We found that concentrations of DD at T2 versus T1 depended on the number of radiofrequency energy applications (r (S) = 0.387; P = 0.002). Marked platelet activation was observed from the start of the procedure, without changes during the procedure.