Data on factor XIII (FXIII) activity in patients on extracorporeal membrane oxygenation (ECMO) are scarce. In this prospective, observational, single-centre study, we determined the rate of acquired FXIII deficiency in 44 adults during ECMO. We evaluated the relationship between minimum FXIII activity, haemoglobin levels, transfusion requirements, as well as the occurrence of major bleeding events. Descriptive statistical methods, bivariate correlation and receiver operating characteristic curve analyses were applied. Acquired FXIII deficiency (FXIII activity < 70%) was found in 41/44 patients [93.2% (95% CI 85.7–100%)] during ECMO. The minimum FXIII activity was correlated with minimum haemoglobin levels (Spearman-rho, 0.331, 95% CI 0.02–0.57, p = 0.03), packed red blood cell transfusion requirements (Spearman-rho, − 0.32, 95% CI − 0.58 to 0.00, p = 0.03), but not the occurrence of major bleeding events (p = 0.32, Hodges-Lehmann difference − 4.0%, 95% CI − 15.0 to 5.0%) during ECMO. The area under the receiver operating characteristic curve for minimum FXIII activity to predict the occurrence of major bleeding events during ECMO was 0.602 (95% CI 0.375–0.828). The lowest FXIII activity to predict a major bleeding event was 51.0% and 55.5% (Youden index for both, 0.27), respectively. In conclusion, acquired FXIII deficiency is highly frequent in adult ECMO patients. Lower FXIII activity showed modest correlations with nadir haemoglobin and pRBC exposure, but did not discriminate patients with major bleeding. Larger studies are needed to determine whether routine FXIII monitoring and targeted replacement improve outcomes during ECMO.