Effect of rejuvenation and frozen storage on 42-day-old AS-1 RBCs

BACKGROUND: The FDA has approved a 42-day storage period for RBCs stored in ADSOL (AS-1). This study was undertaken to provide data for the FDA about the feasibility of salvaging AS-1 RBCs at the end of their storage period by rejuvenation and freezing. STUDY DESIGN AND METHOD: The investigation, consisting of a study (n = 10) and control (n = 6) arm, was carried out in two centers. In both centers, eight healthy volunteers donated a unit (450 mL) of whole blood. The RBC concentrates were stored at 4 degrees C in AS-1 for 42 days. The study units were rejuvenated, whereas the control units were not. All units were stored frozen at -80 degrees C, then deglycerolized and kept for an additional 24 hours at 4 degrees C. RESULTS: After the 42-day storage period, ATP had declined to 62 percent of the original value, 2,3 DPG was zero, and MCV was significantly larger than that of fresh RBCS: Following rejuvenation and deglycerolization, the mean ATP level was 141 percent, the mean 2,3 DPG level was 109 percent, and the MCV was normal. The freeze-thaw-wash recovery of the rejuvenated and nonrejuvenated RBCs was similar, 88.4 and 84.0 percent, respectively. There was no difference in hypoxanthine, inosine, and uric acid levels in the rejuvenated and nonrejuvenated units, which indicated that the chemicals in the rejuvenation solution and their by-products had been removed during processing. In both centers, the mean 24-hour survival of rejuvenated, deglycerolized RBCs exceeded 75 percent, whereas that of nonrejuvenated RBCs did not. The long-term survival rates of viable study and control RBCs were similar. CONCLUSION: Forty-two-day-old AS-1 RBCs that have been rejuvenated and then frozen have more than 75 percent viability and normal oxygen delivery function. Rejuvenation of RBCs does not introduce additional safety hazards to blood transfusion.     

Automation of the glycerolization of red blood cells with the high-separation bowl in the Haemonetics ACP 215 instrument

BACKGROUND: The FDA has approved a closed-system red blood cell (RBC) glycerolization procedure with the ACP 215 (Haemonetics), which requires a centrifuge to prepare RBCs before and after glycerolization. In the study reported here, the Haemonetics high-separation bowl was evaluated in an attempt to automate these two concentration steps. STUDY DESIGN AND METHODS: Ten units of nonleukoreduced citrate phosphate dextrose (CPD)-anticoagulated whole blood were stored at 4 degrees C for 2 to 6 days before glycerolization and freezing as nonrejuvenated RBCs. Twenty-five units of nonleukoreduced CPD whole blood were stored at 4 degrees C for 2 to 8 days and then biochemically treated with a solution containing pyruvate, inosine, phosphate, and adenine (PIPA) before glycerolization and freezing as indated-rejuvenated RBC. Twenty units of leukoreduced CPD and AS-1 RBCs were stored at 4 degrees C for a mean of 48 days and treated with PIPA solution before glycerolization and freezing as outdated-rejuvenated RBCs. The glycerolized RBCs were frozen for at least 2 weeks at -80 degrees C, deglycerolized in the Haemonetics ACP 215 with the 325-mL bowl, and stored in AS-3 at 4 degrees C for 21 days. RESULTS: It took approximately 50 minutes to glycerolize the nonrejuvenated and rejuvenated RBCs. After freezing, deglycerolization, and postwash storage at 4 degrees C in AS-3 for 2 weeks, the quality was similar to that of RBCs processed by the current FDA-approved method. CONCLUSION: Processing time and need for technical expertise were significantly reduced with the completely automated functionally closed glycerolization procedure with the high-separation bowl in the Haemonetics ACP 215 instrument.     

The survival, function, and hemolysis of human RBCs stored at 4 degrees C in additive solution (AS-1, AS-3, or AS-5) for 42 days and then biochemically modified, frozen, thawed, washed, and stored at 4 degrees C in sodium chloride and glucose solution for 24 hours

BACKGROUND: A study was done to assess the quality of RBCs stored at 4 degrees C in AS-1, AS-3, or AS-5 for 42 days before biochemical modification and freezing. STUDY DESIGN AND METHODS: RBCs were stored at 4 degrees C for 42 days in AS-1, AS-3, or AS-5 and then biochemically modified with pyruvate, inosine, phosphate, and adenine solution (Rejuvesol), frozen with 40-percent (wt/vol) glycerol, and stored at -80 degrees C for at least 2 months. The RBCs were deglycerolized by the use of a cell washer (Haemonetics 115), and stored for 24 hours at 4 degrees C in a 0.9-percent sodium chloride and 0.2-percent glucose solution before the autologous transfusion. RESULTS: The mean freeze-thaw-wash recovery process produced RBC recovery values of 85 percent, with the mean 24-hour posttransfusion survival at 75 percent, and the mean index of therapeutic effectiveness at 64 percent for the RBCs stored at 4 degrees C in AS-1, AS-3, or AS-5 for 42 days before biochemical modification and freezing. All the units exhibited normal or slightly higher than normal 2,3 DPG levels after deglycerolization and postwash storage at 4 degrees C for 24 hours. CONCLUSION: RBCs stored in AS-1, AS-3, or AS-5 at 4 degrees C for 42 days and then biochemically modified with pyruvate, inosine, phosphate, and adenine and glycerolized, frozen, washed, and stored at 4 degrees C for 24 hours before autologous transfusion had acceptable in vitro and in vivo measurements.     

Posttransfusion survival of red cells frozen for 8 weeks after 42-day liquid storage in AS-3

Current standards recommend that red cells (RBCs) should be frozen within 6 days of donation. There are situations, however, in which it is desirable to freeze RBCs that are older than 6 days, such as for the salvage of rare or autologous units. To determine the therapeutic efficacy of RBCs frozen after prolonged liquid storage, standard units were drawn from nine normal donors and stored at 4 degrees C for 42 days in a nutrient-additive solution, AS-3. 51CrRBC survival assays were performed (24-hour survival: 78.2 +/- 12.4%; n = 8) and the units were frozen at -80 degrees C in glycerol for 8 weeks. After deglycerolization, the mean RBC recovery was 81.0 +/- 4.1 percent and the mean 24-hour 51Cr survival was 78.0 +/- 9.1 percent. The index of therapeutic effectiveness (ITE) was determined by multiplying the postdeglycerolization 24-hour 51Cr survival by the mean RBC recovery (63.3 +/- 9.2). ITE values greater than 60 percent (75% 51Cr survival x 80% RBC recovery) were considered acceptable. Mean adenosine triphosphate levels declined from an initial 3.81 +/- 0.56 micromol per g of hemoglobin to 2.33 +/- 0.55 micromol per g after frozen storage. These findings show that an acceptable percentage of RBCs survives frozen storage after maximum liquid storage (mean ITE greater than 60%). If necessary, RBCs stored in AS-3 can be frozen at any time before 42 days.     

Posttransfusion survival (24-hour) and hemolysis of previously frozen, deglycerolized RBCs after storage at 4 degrees C for up to 14 days in sodium chloride alone or sodium chloride supplemented with additive solutions

BACKGROUND: Previously frozen human RBCs currently are glycerolized and deglycerolized by the use of open systems that limit storage of the deglycerolized RBCs at 4 degrees C to only 24 hours. STUDY DESIGN AND METHODS: Healthy male volunteers who met AABB requirements for blood donors (n = 38) were studied. A volume of 450 mL of blood was collected into CPDA-1. The RBC concentrates were stored at 4 degrees C for 3 to 6 days before being frozen with 40-percent (wt/vol) glycerol and stored at -80 degrees C. The RBCs were deglycerolized, resuspended in 0.9-percent sodium chloride and 0.2-percent glucose (SG) solution or SG solution supplemented with AS-1, AS-3, or AS-5, and stored in the resuspension medium at 4 degrees C for 14 days. RESULTS: The mean +/- SD freeze-thaw-wash process recovery was 90.0 +/- 4.0 percent for all 38 units. The mean 24-hour posttransfusion survival value was 79 percent for deglycerolized RBC stored at 4 degrees C for 7 days in SG alone, SG plus AS-3, or SG plus AS-5. Deglycerolized RBC that were stored at 4 C for 14 days in SG supplemented with AS-1, AS-3, or AS-5 had a mean 24-hour posttransfusion survival of 74 percent. After 7 days of storage of deglycerolized RBCs in SG alone, the mean hemolysis was 3. 7 percent. After 14 days of storage of deglycerolized RBCs in SG supplemented with AS-1, AS-3, or AS-5, the mean hemolysis was 2.5 percent. CONCLUSIONS: The levels of hemolysis did not correlate with the 24-hour posttransfusion survival values.     

Extended storage of AS-1 and AS-3 leukoreduced red blood cells for 15 days after deglycerolization and resuspension in AS-3 using an automated closed system

BACKGROUND: The utilization of cryopreserved red blood cell (RBC) units had been limited by a maximum postdeglycerolization storage of 24 hours at 1 to 6 degrees C until the recent development of a closed system for the glycerolization and deglycerolization process. STUDY DESIGN AND METHODS: Sixty leukoreduced additive solution (AS), AS-1 (n = 30) and AS-3 (n = 30) RBC units from 500-mL whole blood (WB) collections were stored for 6 days, glycerolized, frozen at -70 +/- 5 degrees C for at least 14 days, thawed, deglycerolized, and stored for 15 days at 1 to 6 degrees C. Glycerolization and deglycerolization were performed with the ACP 215. In-vitro variables were tested before glycerolization, on Day 0, and Day 15 after deglycerolization storage. Forty donors were assessed for double-label 24-hour percent recovery, and T1/2 survival time was measured for 20 donors. RESULTS: Postdeglycerolization mean +/- standard deviation in-vitro RBC mass recoveries were 93 +/- 5 percent for AS-1 and 95 +/- 4 percent for AS-3. Mean hemoglobin +/- standard deviation after deglycerolization was 50.5 +/- 5.5g for AS-1 and 50.1 +/- 3.5g for AS-3. Mean hemolysis (Day 15) was 0.36 +/- 0.11 percent for AS-1 and 0.38 +/- 0.13 percent for AS-3. Double-label 24-hour in-vivo recoveries were 82.5 +/- 7.8 percent for AS-1 and 81.4 +/- 7.1 percent for AS-3. The 51Cr T1/2 value was 41.8 +/- 3.97 for AS-1 and 40.6 +/- 7.11 for AS-3. Other in-vitro variables were as expected. CONCLUSION: Leukoreduced AS-1 and AS-3 RBCs after frozen storage at -70 +/- 5 degrees C can be stored for up to 14 days when processing is performed with the ACP 215 system with resuspension of deglycerolized RBCs in AS-3.     

Evaluation of red blood cells stored at -80 degrees C in excess of 10 years

BACKGROUND: RBCs frozen in 40 percent (wt/vol) glycerol are currently approved by the FDA and the AABB for storage at -80 degrees C for up to 10 years. STUDY DESIGN AND METHODS: This study examined 20 RBC units that had been cryopreserved in 40 percent (wt/vol) glycerol and stored at -80 degrees C for up to 22 years. Measures of the freeze-thaw-wash (FTW) recovery, ATP, 2,3-DPG, methemoglobin, RBC indices, morphology, and osmotic fragility were made immediately after deglycerolization and after 24 hours of storage at 4 degrees C. RESULTS: RBCs frozen for longer than 10 years had acceptable mean FTW recovery, normal oxygen transport function, RBC morphology, RBC indices, methemoglobin, and osmotic fragility. Statistical analysis indicated that the in-vitro viability and function of cryopreserved RBCs was not dependent on the length of frozen storage or postthaw storage at 4 degrees C but did correlate with the storage length at 4 degrees C before cryopreservation. CONCLUSION: The data reported in this study demonstrate that RBCs can be stored at -80 degrees C beyond 10 years with acceptable in-vitro quality and suggest that more defined criteria for the cryopreservation process be adopted.     

In vitro variables of red blood cell components collected by apheresis and frozen 6 and 14 days after collection

BACKGROUND: An automated cell processing system (ACP 215, Haemonetics Corp.) can be used for the glycerolization and deglycerolization of RBC components, but the components must be 6 or fewer days old. Depending on the anticoagulant (CP2D)/additive solution (AS) used, deglycerolized RBCs can be stored at 1 to 6 degrees C for up to 14 days. This study evaluated in vitro variables of apheresis RBC stored for 6 and 14 days at 1 to 6 degrees C before glycerolization and 14 days after deglycerolization. STUDY DESIGN AND METHODS: Two units of CP2D/AS-3 leukoreduced RBCs were collected by apheresis from seven donors. One unit was glycerolized and frozen 6 days and the other 14 days after collection. All units were deglycerolized with the ACP 215 and stored at 1 to 6 degrees C for 14 days in AS-3. Several in vitro variables were evaluated during postdeglycerolization storage. RESULTS: All components had postdeglycerolization RBC recoveries greater than 81 percent and osmolalities of less than 400 mOsm per kg. No significant differences were noted in potassium and supernatant hemoglobin after 14 days of postdeglycerolization storage between RBCs frozen at 6 and 14 days after collection. After 14 days of postdeglycerolization storage, however, the pH, lactate, and ATP levels were slightly lower in RBCs frozen after 14 days. CONCLUSION: The ACP 215 can be used to glycerolize and deglycerolize apheresis RBC components that are up to 14 days of age. It is likely that apheresis components glycerolized at 14 days of age or less can be stored up to 14 days in AS-3 after deglycerolization, but this should be confirmed with in vivo survival studies.     

A multicenter study of in vitro and in vivo values in human RBCs frozen with 40-percent (wt/vol) glycerol and stored after deglycerolization for 15 days at 4°C in AS-3: assessment of RBC processing in the ACP 215

BACKGROUND: The FDA has approved the storage of frozen RBCs at -80 degrees C for 10 years. After deglycerolization, the RBCs can be stored at 4 degrees C for no more than 24 hours, because open systems are currently being used. Five laboratories have been evaluating an automated, functionally closed system (ACP 215, Haemonetics) for both the glycerolization and deglycerolization processes. STUDY DESIGN AND METHODS: Studies were performed at three military sites and two civilian sites. Each site performed in vitro testing of 20 units of RBCs. In addition, one military site and two civilian sites conducted autologous transfusion studies on ten units of previously frozen, deglycerolized RBCs that had been stored at 4 degrees C in AS-3 for 15 days. At one of the civilian sites, 10 volunteers received autologous transfusions on two occasions in a randomized manner, once with previously frozen RBCs that had been stored at 4 degrees C in AS-3 for 15 days after deglycerolization and once with liquid-preserved RBCs that had been stored at 4 degrees C in AS-1 for 42 days. RESULTS: The mean +/- SD in vitro freeze-thaw-wash recovery value was 87 +/- 5 percent; the mean +/- SD supernatant osmolality on the day of deglycerolization was 297 +/- 5 mOsm per kg of H(2)O, and the mean +/- SD percentage of hemolysis after storage at 4 degrees C in AS-3 for 15 days was 0.60 +/- 0.2 percent. The paired data from the study of 10 persons at the civilian site showed a mean +/- SD 24-hour posttransfusion survival of 76 +/- 6 percent for RBCs that had been stored at 4 degrees C for 15 days after deglycerolization and 72 +/- 5 percent for RBCs stored at 4 degrees C in AS-1 for 42 days. At the three sites at which 24-hour posttransfusion survival values were measured by three double-label procedures, a mean +/- SD 24-hour posttransfusion survival of 77 +/- 9 percent was observed for 36 autologous transfusions to 12 females and 24 males of previously frozen RBCs that had been stored at 4 degrees C in AS-3 for 15 days after deglycerolization. CONCLUSION: The multicenter study showed the acceptable quality of RBCs that were glycerolized and deglycerolized in the automated ACP 215 instrument and stored in AS-3 at 4 degrees C for 15 days.     

In vivo survival of apheresis RBCs, frozen with 40-percent (wt/vol) glycerol, deglycerolized in the ACP 215, and stored at 4°C in AS-3 for up to 21 days

BACKGROUND: The FDA has approved the storage of frozen RBCs at -80 degrees C for 10 years and the postwash storage at 4 degrees C for no more than 24 hours. The 4 degrees C postwash storage period is limited to 24 hours, because the current deglycerolization systems are functionally open systems. STUDY DESIGN AND METHODS: Two units of RBCs were collected from each of 13 healthy male volunteers. The RBCs were collected in CP2D by the FDA-approved protocol for an automated apheresis device (MCS, LN8150, Haemonetics) and were stored at 4 degrees C in AS-3 for 6 days. Using a single disposable glycerolization set in an automated, functionally closed system (ACP 215, Haemonetics) each unit was transferred to a 1000-mL PVC plastic bag and glycerolized to a concentration of 40-percent (wt/vol) glycerol and frozen at -80 degrees C. A single disposable deglycerolization set in the ACP 215 was used to deglycerolize the 2 units from the same donor. The deglycerolized RBCs were stored at 4 degrees C in AS-3 for as long as 21 days. RESULTS: The mean +/- SD freeze-thaw-wash recovery value was 89.4 +/- 3 percent. The residual hemolysis in the RBCs stored at 4 degrees C in AS-3 for 21 days after deglycerolization was 0.9 +/- 0.2 percent, and the units were negative for both aerobic and anaerobic bacteria. The mean Nageotte WBC count was 9 x 10(6) per unit. When the deglycerolized RBCs were given as an autologous transfusion after storage at 4 degrees C in AS-3 for the 7- to 18-day period, the mean +/- SD 24-hour posttransfusion survival was 77 +/- 7 percent, and the index of therapeutic effectiveness was 69 +/- 8 percent. CONCLUSION: Two units of human RBCs collected from a single donor by apheresis in the MCS using an LN8150 set can be glycerolized sequentially with a single disposable set and deglycerolized sequentially with another single disposable set in the ACP 215. The previously frozen RBCs stored in AS-3 for 7 to 18 days at 4 degrees C had acceptable hemolysis and an acceptable mean 24-hour posttransfusion survival value and index of therapeutic effectiveness.     

Altered processing of thawed red cells to improve the in vitro quality during postthaw storage at 4 degrees C

BACKGROUND: The use of a functionally closed system (ACP215, Haemonetics) for the glycerolization and deglycerolization of red blood cell (RBC) units allows for prolonged postthaw storage. In this study, the postthaw quality of previously frozen, deglycerolized RBCs resuspended in saline-adenine-glucose-mannitol (SAGM) or additive solution AS-3 was investigated. STUDY DESIGN AND METHODS: Leukoreduced RBC units were frozen with 40 percent glycerol and stored at -80 degrees C for at least 14 days. The thawed units were deglycerolized with the ACP215, resuspended in SAGM or AS-3, and stored at 2 to 6 degrees C for up to 21 days. RESULTS: The mean +/- standard deviation in vitro freeze-thaw-wash recovery was 81 +/- 5 percent. During storage, hemolysis of deglycerolized cells remained below 0.8 percent for 2 days in SAGM and for 14 days in AS-3. This difference was explained by the protective effect of citrate, which is present in AS-3. Cells stored in AS-3 showed a lower glycolytic activity and a faster decline in adenosine 5'-triphosphate (ATP) than cells in SAGM. Increasing the internal pH of cells before storage in AS-3 by use of phosphate-buffered saline (PBS) in the deglycerolization procedure resulted in elevated lactate production and better maintenance of intracellular ATP content. After 3 weeks of storage, the ATP content of PBS-washed cells amounted to 2.5 +/- 0.5 micromol per g of hemoglobin (Hb), whereas for saline/glucose-washed cells this value was decreased to 1.0 +/- 0.3 micromol per g of Hb. CONCLUSIONS: Leukoreduced, deglycerolized RBCs can be stored for 48 hours in SAGM. Improved ATP levels during refrigerated storage can be observed with thawed cells, resuspended in AS-3, when PBS is used as a washing solution.     

Automated collection of double red blood cell units with a variable-volume separation chamber

BACKGROUND: Automated collection of blood components offers multiple advantages and has prompted development of portable devices. This study sought to document the biochemical and hematologic properties and in vivo recovery of red cells (RBCs) collected via a new device that employed a variable-volume centrifugal separation chamber. STUDY DESIGN AND METHODS: Normal subjects (n = 153) donated 2 units of RBCs via an automated blood collection system (Cymbal, Haemonetics). Procedures were conducted with wall outlet power (n = 49) or the device's battery source (n = 104). Units were collected with or without leukoreduction filtration and were stored in AS-3 for 42 days. The units were assessed via standard biochemical and hematologic tests before and after storage, and 24 leukoreduced (LR) and 24 non-LR RBCs were radiolabeled on Day 42 with Na(2)(51)CrO(4) for autologous return to determine recovery at 24 hours with concomitant determination of RBC volume via infusion of (99m)Tc-labeled fresh RBCs. RESULTS: Two standard RBC units (targeted to contain 180 mL of RBCs plus 100 mL of AS-3) could be collected in 35.7 +/- 2.0 minutes (n = 30) or 40.3 +/- 2.7 minutes for LR RBCs (n = 92). An additional 31 collections were conducted successfully with intentional filter bypassing. RBC units contained 104 +/- 4.1 percent of their targeted volumes (170-204 mL of RBCs), and LR RBCs contained 92 percent of non-LR RBCs' hemoglobin. All LR RBCs contained less than 1 x 10(6) white blood cells. Mean hemolysis was below 0.8 percent (Day 42) for all configurations. Adenosine triphosphate was well preserved. Mean recovery was 82 +/- 4.9 percent for RBCs and 84 +/- 7.0 percent for LR RBCs. CONCLUSIONS: The Cymbal device provided quick and efficient collection of 2 RBC units with properties meeting regulatory requirements and consistent with good clinical utility.     

A hypotonic storage solution did not prolong the viability of RBCs

BACKGROUND: Hypotonic storage solutions and WBC filtration are both reported to improve RBC viability. This study tested the ability of an investigational hypotonic storage solution (AS-24, Medsep Corp.) to extend the viability of liquid-stored RBCs to 8 weeks. STUDY DESIGN AND METHODS: In a pair of crossover trials, 11 RBC units, WBC-reduced by filtration and stored in AS-24 for 8 weeks, were compared with units from the same donors that were stored for 6 weeks in AS-3, and 13 RBC units, WBC-reduced by filtration and stored in AS-3 for 8 weeks, were compared with units from the same donors that were stored for 6 weeks in AS-3. Viability was measured by the (51)Cr/(99m)Tc double-isotope method. RESULTS: RBC viability at 8 weeks averaged 64 +/- 3 percent in the AS-24 units and 67 +/- 2 percent in the AS-3 units. It was equal at 77 +/- 3 percent and 77 +/- 2 percent after 6 weeks' storage in AS-3 in both trials. CONCLUSIONS: Prestorage WBC reduction and storage in AS-24 did not extend RBC viability to 8 weeks. The improved viability previously demonstrated with storage of dilute suspensions of RBCs in hypotonic solutions is probably caused by factors other than the hypotonicity.     

The viability of autologous human red cells stored in additive solution 5 and exposed to 25°C for 24 hours

BACKGROUND: No data exist on the viability of red cells (RBCs) stored in modern additive solution systems and allowed to warm above 10 degrees C. STUDY DESIGN AND METHODS: In a randomized crossover study, 3 units of blood were collected at least 8 weeks apart from 11 volunteer donors and stored in additive solution 5 (AS-5). Of 3 units from each volunteer, 1 was stored for 6 weeks at 4 degrees C, 1 for 5 weeks at 4 degrees C except for 24 hours at 25 degrees C on Day 14, and 1 for 5 weeks at 4 degrees C except for 24 hours at 25 degrees C on Day 28. Units were sampled periodically during storage; at the end of storage, viability was measured by the 99mTc/51 Cr double-label method. RESULTS: RBC viability was not significantly different in the storage protocols. Less than 1 percent of stored cells hemolyzed. RBC ATP concentrations at the end of storage correlated with viability and were approximately equal in the warmed units after 30 days' storage and the conventionally stored units after 42 days. CONCLUSIONS: The data suggest that RBCs stored in AS-5 and allowed to warm to 25 degrees C for 24 hours lose about 12 days of their shelf life.     

Addition of L-carnitine to additive solution-suspended red cells stored at 4 degrees C reduces in vitro hemolysis and improves in vivo viability

BACKGROUND: The role of L-carnitine (LC) as the requisite carrier of long-chain fatty acids into mitochondria is well established. Human red cells (RBCs), which lack mitochondria, possess a substantial amount of LC and its esters. In addition, carnitine palmitoyl transferase, an enzyme that catalyzes the reversible transfer of the acyl moiety from acyl-coenzyme A to LC is found in RBCs. It has recently been shown that LC and carnitine palmitoyl transferase play a major role in modulating the pathway for the turnover of membrane phospholipid fatty acids in intact human RBCs, and that LC improved the membrane stability of RBCs subjected to high shear stress. RBC membrane lesions occur during storage at 4 degrees C; this study investigated whether the addition of LC (5 mM) to a standard RBC preservative solution (AS-3) affected cellular integrity with 42 days' storage. STUDY DESIGN AND METHODS: A paired (n = 10) crossover design was used for RBCs stored in AS-3 with and without LC. Both in vitro RBC properties reflective of metabolic and membrane integrity and in vivo measures of cell viability (24-hour percentage of recovery and circulating lifespan) were measured at the end of the storage. In addition, the turnover of membrane phospholipid and long-chain acylcarnitine fatty acids and the carnitine content of control and LC-stored RBCs were measured. RESULTS: It was shown that LC was irreversibly taken up by RBCs during storage, with a fourfold increase at 42 days. Furthermore, as found by the use of radiolabeled palmitate, the stored RBCs were capable of generating long-chain acylcarnitine. The uptake of LC during storage was associated with less hemolysis and higher RBC ATP levels and by a significantly greater in vivo viability for LC-stored RBCs than for control-stored RBCs: a mean 24-hour percentage of recovery of 83.9 +/? 5.0 vs. 80.1 +/? 6.0 percent and a mean lifespan of 96 +/? 11 vs. 86 +/? 14 days, respectively (p < 0.05). CONCLUSION: A beneficial effect of the addition of LC to RBCs stored at 4 degrees C was evident. This effect may be related to both biophysical and metabolic actions on the cell membrane.     

Anaerobic storage of red blood cells in a novel additive solution improves in vivo recovery

BACKGROUND: In preliminary studies, anaerobic red blood cell (RBC) storage reduced oxidative damage and phosphatidylserine exposure while maintaining adenosine triphosphate levels. The purpose of this study was to compare the 24-hour recovery and life span of autologous RBCs stored 6 and 9 weeks using OFAS3 additive solution in an anaerobic environment, compared to control RBCs aerobically stored in AS-3 for 6 weeks.
STUDY DESIGN AND METHODS: Eight subjects were entered into a randomized, crossover study. Whole blood was collected from each subject twice separated by 12 weeks or more into CP2D and leukoreduced. Controls were stored in AS-3. Test units in OFAS3 were oxygen depleted with argon then stored 9 weeks in an anaerobic chamber at 1 to 6°C. At the end of each storage period, RBCs were labeled with ⁵¹Cr and ^99mTc and reinfused to the subject following standard methods to determine double-label recovery and life span. Hypotheses tests were conducted using paired, repeated-measures analysis of variance.
RESULTS: Recovery for the anaerobically stored test RBC was significantly better than control at 6 weeks (p = 0.023). Test units at 9 weeks were not different than the 6-week control units (p = 0.73). Other in vitro measures of RBC characteristics followed the same trend. Two test units at 9 weeks had hemolysis of greater than 1 percent.
CONCLUSION: Anaerobically stored RBCs in OFAS3 have superior recovery at 6 weeks compared to the controls and equivalent recovery at 9 weeks with no change in life span. Anaerobic storage of RBCs may provide improved RBCs for transfusion at 6 weeks of storage and may enable extending storage beyond the current 42-day limit.     

Rejuvenation of irradiated AS-1 red cells

BACKGROUND: Gamma irradiation of blood components is used to prevent transfusion-associated graft-versus-host disease. The demand for irradiated blood components is increasing because of the increase in directed donation by family members. Irradiated units currently have a recommended maximum storage life of 28 days. Since in vivo recovery is related to red cell ATP levels, rejuvenation of stored irradiated units using a pyruvate-inosine phosphate-adenine additive was explored. STUDY DESIGN AND METHODS: Units of AS-1 red cells from 16 volunteer donors were divided into two equal volumes and one split unit from each was irradiated with 25 Gy. Ten units were irradiated on Day 5, 6, or 7 of 4 degrees C storage and 6 units were irradiated on Day 1 of 4 degrees C storage. All units were rejuvenated for 1 hour at 37 degrees C using a pyruvate-inosine-phosphate-adenine additive on Day 42 of 4 degrees C storage. Units were assayed for ATP, 2, 3 DPG and supernatant sodium, potassium, and glucose. RESULTS: ATP and 2, 3 DPG levels were restored equally well in irradiated and non-irradiated units. The previously reported irradiation-induced red cell potassium-sodium shift was demonstrated. Supernatant potassium and sodium levels did not reverse 1 hour after rejuvenation was completed. There was no significant difference in results between units irradiated on Day 1 or Day 5, 6, or 7. CONCLUSION: Red cell ATP and 2, 3 DPG levels were restored in irradiated AS-1 units stored at 4 degrees C for 42 days using a pyruvate-inosine-phosphate-adenine rejuvenation additive.     

The in vitro quality of red blood cells frozen with 40 percent (wt/vol) glycerol at -80 degrees C for 14 years, deglycerolized with the Haemonetics ACP 215, and stored at 4 degrees C in additive solution-1 or additive solution-3 for up to 3 weeks

BACKGROUND: Red blood cells (RBCs) frozen with 40 percent (wt/vol) glycerol, stored at -80 degrees C (mean temperature; range, -65 to -90 degrees C) for 14 years, deglycerolized in the Haemonetics automated cell processor (ACP) 215 with the 325-mL disposable bowl, and stored at 4 degrees C in additive solution (AS)-1 or AS-3 for 21 days were evaluated. STUDY DESIGN AND METHODS: A total of 106 units of citrate phosphate dextrose adenine-1 RBCs were frozen with 40 percent (wt/vol) glycerol in the original 800-mL polyvinylchloride plastic bag and stored in corrugated cardboard boxes at -80 degrees C for 14 years. The thawed units were deglycerolized with the ACP 215 with a 325-mL disposable bowl and stored in AS-1 or AS-3 at 4 degrees C for 21 days. RESULTS: The freeze-thaw recovery value was 94 +/- 4 percent (mean +/- SD), the freeze-thaw-wash recovery value was 80 +/- 7 percent, and there was no breakage. Thirty-eight units were processed as 19 pairs. Two units of ABO-matched units were thawed, pooled, divided equally into two units, and deglycerolized. One unit was stored in AS-1 and the other in AS-3 at 4 degrees C for 21 days. Units stored in AS-1 exhibited significantly greater hemolysis than those stored in AS-3. CONCLUSIONS: Acceptable results were achieved when RBCs frozen at -80 degrees C for 14 years were deglycerolized in the ACP 215. Deglycerolized RBCs in AS-1 exhibited significantly higher hemolysis than those in AS-3 after storage at 4 degrees C for 7 to 21 days.     

The 24-hour posttransfusion survival of baboon red blood cells preserved in citrate phosphate dextrose/ ADSOL (CPD/AS-1) for 49 days.

The purpose of this study was to evaluate the baboon as an animal model for evaluating red blood cell (RBC) preservation by comparing the 24-h posttransfusion survival of baboon RBCs preserved in citrate phosphate dextrose/ADSOL (CPD/AS-1) solution at 4 degrees C for 49 days to that of human RBCs preserved under similar conditions. CPD/AS-1 originally was approved by the Food and Drug Administration for 49-day storage of RBCs, but this period subsequently was reduced to 42 days. Adult male baboons (Papio anubis and P. cynocephalus) were autotransfused with RBCs that had been harvested using CPD and that had been resuspended and stored in AS-1 solution at 4 degrees C for as long as 49 days. The 24-h posttransfusion survival was measured using the 51Cr/125I-albumin method. The 24-h posttransfusion survival (mean +/- standard deviation) was 74% +/- 7% for seven units of CPD/AS-1-treated RBCs stored for 35 days, 65% +/- 15% for 12 units stored for 42 days, and 43% +/- 16% for seven units stored for 49 days. The mean 24-h posttransfusion survival rate for autologous baboon RBCs stored in CPD/AS-1 at 4 degrees C for 35 days (74%) was similar to that for autologous human RBCs stored in a similar manner. Further storage for 42 and 49 days resulted in lower values for baboon RBCs compared with human RBCs.     

A feasibility evaluation of an automated bloodcomponent collection system platelets and red cells

BACKGROUND: The purpose of these studies was to evaluate the functional properties of blood components collected with an automated collection system. STUDY DESIGN AND METHODS: Single-donor platelets (n = 44) and packed red cell (RBC) units (n = 10) were collected. In vitro and in vivo assays were used to assess the function of single-donor platelet components stored for 5 days and of packed RBC units after storage for 42 days at 4 degrees C. RESULTS: Adverse events observed in the 44 study subjects were minor. The mean 24-hour recovery value for the packed RBC units stored for 42 days was 83.6 +/- 5.4 percent, with a mean percentage of hemolysis on Day 42 at 0.46 +/- 0.19 percent. The 25 patients receiving platelet components achieved a mean corrected count increment of 15.1 +/- 10.4 x 10(3). All platelet concentrates had less than 1 x 10(6) total white cells. CONCLUSION: Both in vitro and in vivo testing for the packed RBCs collected and stored for 42 days met the standards for both hemolysis and percentage of 51Cr 24-hour RBC recovery. The in vitro results and transfusion data on white cell-reduced platelet components transfused to thrombocytopenic patients were comparable to those on available platelet components.