Manufacturing method affects mitochondrial DNA release and extracellular vesicle composition in stored red blood cells

Background and Objectives Damage‐associated molecular patterns ( DAMP s) are found in transfusion products, but their potential impacts are not fully understood. We examined the influence of manufacturing method on levels of mitochondrial (mt) DNA and extracellular vesicle ( EV ) DAMP s in red cell concentrates ( RCC s). Materials and Methods Eighty‐seven RCC s were prepared using nine different methods (6–15 units/method), including three apheresis, five whole blood ( WB )‐derived leucoreduced ( LR ) and one WB ‐derived non‐ LR method. On storage days 5 and 42, levels of mt DNA (by PCR ) and number and cell of origin of EV s (by flow cytometry) were assessed in RCC supernatants. Results There was a 100‐fold difference in mt DNA levels among methods, with highest levels in non‐ LR , followed by MCS + and Trima apheresis RCC s. There was a 10‐fold difference in EV levels among methods. RBC ‐derived CD 235a+ EV s were found in fresh RCC s and increased in most during storage. Platelet‐derived CD 41a+ EV s were highest in non‐ LR and Trima RCC s and did not change during storage. WBC ‐derived EV s were low in most RCC s; CD 14+ EV s increased in several RCC s during storage. Conclusion DAMP s in RCC s vary by manufacturing method. Mt DNA and EV could be informative quality markers that may be relevant to RCC immunomodulatory potential.