Prolonged storage of purified granulocyte concentrates: Introduction of a new purification method

Abstract

Background Use of donor granulocyte concentrate (GC) has been limited due to its short storage time of 6–24 h, which is partially due to residual red blood cells (RBCs) and platelets and the resulting lactate production leading to an acidotic milieu. To increase this storage time, we developed a closed system procedure compatible with standard blood bank technologies to remove RBC and platelets and to enrich the GC. Methods Standard GCs (sGCs) were sedimented, washed twice with 0.9% sodium chloride (NaCl), and resuspended in blood group‐identical fresh frozen plasma. The resulting purified GCs (pGCs) were then stored in platelet bags at a cell concentration of about 5 × 107 ± 1.8 × 107 leukocytes/ml without agitation at room temperature for up to 72 h. Cell count and viability, pH, blood gases, phagocytosis, and oxidative burst were monitored daily. Results A significant reduction in RBC (98%) through sedimentation, and platelets (96%) by washing, purified the white blood cell (WBC) population and enriched the granulocytes to 96% of the WBC in the pGC. After 72 h of storage, over 90% of the initial WBC count of pGC remained, was viable (≥97%), and the granulocytes exhibited a high phagocytosis and oxidative burst functionality, comparable to sGC after 24 h. Conclusion Purification extends the maximum storage period of GC from 24 to 72 h and may therefore improve the availability of GC and its clinical use.

Extracorporeal therapy of sepsis by purified granulocyte concentrates: ex vivo circulation model

“…Incorporation of special storage bags designed to release the carbon dioxide produced by the cells during storage further contributes to improved pH values of the purified GC supplied to the circuit. 18 In contrast, lactate concentration of septic plasma was elevated in our experiments. This is in correspondence to the disease-specific metabolic environment of sepsis patients.…”

“…The major reason for this is the significantly reduced erythrocyte and platelet concentration of purified GC due to the purification process. 18 In 3 days, stored purified GC and, especially in sepsis plasma circuits, lactate levels are elevated. However, these remain lower when compared to standard GC.…”

“…Hence, a dose of approximately 1.5 × 10 10 cells being targeted and, on top of that, further development of GC was pursued in order to improve the storability and availability of preparations. 18 Use of purified GC in the circuit showed preservation of oxygen and carbon dioxide concentration without the need to implement an oxygenator as an additional device. Furthermore, purification of the standard GC resulted in lower carbon dioxide production thus, system design allowed further simplification for a more practical application on the patient.…”

“…The oxygenator led to an unfavorable complexity of the system and thus significantly limits practical applicability. Hence, a dose of approximately 1.5 × 10 10 cells being targeted and, on top of that, further development of GC was pursued in order to improve the storability and availability of preparations 18 . Use of purified GC in the circuit showed preservation of oxygen and carbon dioxide concentration without the need to implement an oxygenator as an additional device.…”

“…Development of the purification process and the successful extension of storage time to at least 3 days have been described recently. 18 In short, standard GC was sedimented to remove RBC, washed twice with 0.9% saline solution to remove platelets, split into units containing about 1.5 × 10 10 granulocytes, and stored in ABO-compatible, citrate anticoagulated blood plasma with glucose in gas permeable storage bag. The purification procedure of sedimentation and washing reduced the RBC and platelet counts in purified GC, by approximately 98.08 ± 1.87% and 95.91 ± 5.1%, respectively, compared with standard GC.…”