Cost comparison of methods for preparation of neonatal red cell aliquots.

OBJECTIVE: The purpose of this study was to compare the preparation costs of two common methods used for neonatal red blood cell transfusion aliquots. METHODS: Three months of data from a Level 2 and Level 3 neonatal intensive care unit (NICU) were used to determine the comparative cost for red cell aliquot transfusions using an eight bag aliquot/transfer system or the syringe set system. Using leuko-poor red blood cell blood collected in Adsol and containing approximately 320 ml of red blood cells and supernatant solution, the average cost of neonatal transfusion aliquots was determined using the Charter Medical syringe set and the Charter Medical eight bag aliquot/transfer system. RESULTS: A total of 126 red blood cell transfusion aliquots were used over the three month period. The amount transfused with each aliquot ranged from 5.0 ml - 55.0 ml with an average of 24.0 ml per aliquot. The cost per aliquot using the eight aliquot/transfer set was calculated as $36.25 and the cost per aliquot using the syringe set cost was calculated as $30.71. Additional benefits observed with the syringe set included decreased blood waste. CONCLUSION: When comparing Charter Medical multiple aliquot bag sets and the Charter Medical syringe aliquot system to provide neonatal transfusions, the use of the syringe system decreased blood waste and proved more cost effective.     

Storage of Erythrocytes in Artificial Media

The storage of red blood cells (RBC) for extended periods in artificial media without plasma has been studied. Blood was collected in either heparin or ACD solution; the plasma was removed; and one or two volumes of a solution containing 2 to 3 mM adenine, 5 to 60 mM Na₂HPO₄ 55 mM glucose, and 120 to 140 mM NaCl was added to the packed RBC. Control samples were stored in ACD plasma containing an equivalent concentration of adenine (ACD-ad). Viability studies were done on three consecutive days in each of 22 subjects, using the subject's own blood stored in various preservatives for 41 to 57 days. After this period of storage, the mean viability of ACD-ad stored blood was 73.5 per cent and of erythrocytes stored in artificial media, 74.4 per cent. Cells stored for longer periods had diminished viability, but the viability of cells stored in artificial media was equivalent to or superior to that of controls. After 42 days' storage, 2,3 DPG levels of RBC stored in artificial media were higher than those of controls, and in some instances, the 2,3 DPG content was one-third to two-thirds that of fresh blood. Some of the potential advantages of this system for blood preservation are:
1. The suspending medium is discarded prior to infusion so that less potentially toxic substances are administered.
2. Plasma is removed at the beginning of storage so that labile factors are available for fractionation.
3. 2,3 DPG levels are higher, so that, theoretically, the oxygen-delivering capacity of the transfused cells is greater.     

Hemoglobin Function of Blood Stored at 4 C in ACD and CPD with Adenine and Inosine

Normal hemoglobin function depends on adequate erythrocyte levels of 2,3‐diphosphoglycerate (2,3‐DPG), a compound that is poorly maintained during blood bank storage in acid‐citrate‐dextrose (ACD). Since 2,3‐DPG is better maintained at the higher pH afforded by citrate‐phosphate‐dextrose (CPD), hemoglobin function was compared during storage in CPD and ACD. Further, hemoglobin function was studied in CPD blood containing adenine and inosine, compounds that provide metabolic energy and thus prolong the shelf‐life of blood, because they also effect the levels of 2,3‐DPG during storage. Hemoglobin function, expressed as the P₅₀ (the P₀₂ at 50 per cent oxygenation, an inverse but direct measure of oxygen affinity) is considerably better maintained during storage in CPD than in ACD. The hemoglobin function or P₅₀ of blood stored in CPD‐adenine is not maintained as well as blood stored in CPD without adenine, but the oxyhemoglobin dissociation curves show only a small difference when compared to the difference between ACD and CPD. Blood stored in CPD‐adenine with inosine, present initially or added at day 25, allows higher P₅₀ values late in storage, thus providing better hemoglobin function for more of the storage period.     

Characteristics of Recently Outdated ACD Blood and Some Determinants of these Characteristics

Twenty samples of outdated ACD blood were characterized by the following determinations: nucleotide composition, osmotic fragility, p H, hematocrit, final glucose concentration, plasma hemoglobin, and plasma K⁺. In all tests, considerable variability existed among samples. In four samples, these determinations were made daily from day 21 to day 25. In this series there were no consistent daily trends, variability being greater from sample to sample than from day to day. In another study, six stored blood samples from apparently healthy young adults were followed weekly for ATP. Those samples that initially had higher ATP levels retained this differential throughout storage, the rate of ATP breakdown being nearly the same for all samples. Of these samples, those with higher blood ATP levels had faster glycolytic rates.     

Some in Vitro Effects of Adenine Added to Stored Blood

When adenine was added to freshly collected ACD blood and the blood incubated at 37 C. for one or two days, the concentration of ATP and adenine nucleotides (AMP + ADP + ATP) was higher than in the same blood without adenine. The addition of inosine with adenine was even more effective. Adenine was used by the red cells and the effective amounts of adenine to be added varied with the experimental conditions, for example, length of incubation. When adenine or adenine and inosine were incubated with outdated ACD blood, the ATP and adenine nucleotide concentrations rose, indicating that synthetic abilities were still present in red cells which had deteriorated energetically as a result of storage in citrate in the cold. When outdated red cells were returned to a more normal p H, the ATP levels increased. When adenine addition was included along with neutralization, the effects were remarkable, the ATP and adenine nucleotide levels going even above normal fresh blood levels. These experiments suggest that added adenine enhances ATP synthesis and allows formulation of a scheme for the participation of adenine in the metabolic reactions of stored cells.     

Volume of RBCs, 24- and 48-hour posttransfusion survivals, and the lifespan of (51)Cr and biotin-X-N-hydroxysuccinimide (NHS)-labeled autologous baboon RBCs: effect of the anticoagulant and blood pH on (51)Cr and biotin-X-NHS elution in vivo

BACKGROUND: The RBC injury that occurs during collection of the first few milliliters of blood into the pH 5.0 ACD (NIH, Formula A) is referred to as the lesion of collection. The RBC injury was evaluated by labeling the ACD RBCs with (51)Cr and measuring the 24-hour posttransfusion survival. The effect of the acidification of ACD blood on the in vivo elution of (51)Cr from the RBC has not been reported. STUDY DESIGN AND METHODS: Baboon blood was collected in heparin and in ACD and CP2D at different ratios of blood to anticoagulant. ACD blood with a pH of 5.7 to 6.9 was labeled with (51)Cr. Heparinized blood with a pH of 7.4 was labeled with biotin-X-N-hydroxysuccinimide (NHS). ACD blood with a pH of 5.9 was labeled with both (51)Cr and biotin-X-NHS. The RBC volumes, 24- and 48-hour posttransfusion survivals, and lifespans (T50) were measured. RESULTS: The RBC volume of ACD blood with a pH ranging from 5.7 to 6.9 was not affected by (51)Cr labeling. (51)Cr-labeled ACD blood with a pH of 6.9 had an RBC volume that was significantly greater than that seen in heparinized blood with a pH of 7.4 labeled with biotin-X-NHS. In vivo elution of (51)Cr from the RBCs prepared from the ACD blood with a pH of 5.7 to 6.9 and in vivo elution of biotin-X-NHS from RBCs prepared from ACD blood with a pH of 5.9 were associated with reductions in 24- and 48-hour posttransfusion survivals and T50. CONCLUSIONS: The anticoagulant and the pH of the medium in which the RBCs were labeled with (51)Cr or biotin-X-NHS affected in vivo elution of the label from the RBCs and may reduce posttransfusion survival values.     

Albumin and Hydroxy‐Ethyl Starch in the Cryopreservation of Red Cells—an In Vitro Study

The usefulness of a fully dialyzed albumin in ACD‐saline containing hyroxyethyl starch (HES) for cryoprotection of red blood cells has been investigated. In vitro damage was as assessed by the amount of hemolysis in the supernatant plasma, saline stability, and autohemolysis at 30 C after resuspension in fresh ACD plasma. Freeze‐thawing hemolysis was reduced to an average of 54 per cent of that of the control when the cells were processed in 14.1 g 100 ml albumin containing 11.7 g 100 ml HES. Higher concentrations of albumin reduced this difference of 46 per cent to 20 per cent. The autohemolysis test gave the best result, when 21.7 g/100 ml albumin was used as HES diluent.
Additionally the effect of freeze‐thawing on the cellular levels of ATP and 2,3‐DPG was investigated. The ATP levels in the albumin‐HES processed cells fell to 94 per cent and that of the ACD plasma‐HES cells to 85 per cent of the respective starting values. 2,3‐DPG in the corresponding samples was reduced to 98 and 96 per cent of the initial levels. The differences between cells processed in both media were statistically not significant. Cell membrane deformability was tested by cell filtration. Cells processed in albumin‐HES had a shorter filtration time than those frozen in ACD plasma‐HES and this difference was statistically significant.     

Serum transferrin receptor assay in iron deficiency anaemia and anaemia of chronic disease in the elderly

The most common cause of anaemia in the elderly is anaemia of chronic disease (ACD). However, iron deficiency anaemia (IDA) may coexist, and can be difficult to diagnose. The serum transferrin receptor (sTfR) blood test may be a better indicator of iron status as it is not affected by inflammation nor by advancing age. We evaluated it in four groups (10 males, 10 females each): 'young' controls, 'elderly' controls, IDA and ACD. All patients in the IDA group had elevated sTfR levels (mean +/- SD 65.2 +/- 17.7 nmol/l). All 'young' controls had normal sTfR (22.3 +/- 7.3 nmol/l) and ferritin levels (92.7 +/- 61.1 micrograms/l). Although all subjects in the 'elderly' controls and ACD group had normal, and raised or normal serum ferritin, respectively (88 +/- 62.3 micrograms/l; 631.2 +/- 509.5 micrograms/l), three (15%) 'elderly' controls and four (20%) ACD patients had raised sTfR levels, suggesting depleted iron stores. Bone-marrow aspirates were available in 3/4 ACD patients with raised sTfR. Haemosiderin was absent in two. The sTfR blood test is comparable to serum ferritin in diagnosing IDA in the elderly but also seems capable of differentiating ACD from IDA. Its potential as a non-invasive test of iron status, especially in elderly anaemic patients, deserves further evaluation.     

自体血贮存在不同温度时红细胞形态的变化

背景：输血指南指出：全血应在（4±2）℃贮存,血液一旦离开正确的贮存条件,即有发生细菌繁殖或丧失功能的危险,受血者输注后则会发生不同程度的输血反应或输注无效。 目地：利用显微镜观察自体血贮存在不同温度下红细胞形态的变化。 方法：40例急性等容血液稀释患者取血后贮存在ACD枸橼酸血袋,分别贮存在4 ℃和常温23 ℃,于放自体血后即刻、自体血贮存1,2,3,4,5,6 h分别取样涂片,利用显微镜观察红细胞形态变化,计算红细胞畸形率。随机选取6 h段常温组血样与有效期内的ACD库存血各6份进行pH、K＋、游离血红蛋白等对比及细菌培养。 结果与结论：4 ℃组和常温组在各时间点红细胞畸形率差异无显著性意义, 6 h段常温组血样pH、K＋、游离血红蛋白等均优于有效期内的ACD库存血,培养均无细菌生长。提示常温下自体血贮存6 h内回输给患者是可行的。     

Quantitation of genomic DNA in plasma and serum samples: higher concentrations of genomic DNA found in serum than in plasma

BACKGROUND: Plasma and serum samples have been used to detect cell-free genomic DNA in serum or plasma in certain pathologic conditions such as systemic lupus erythematosus, pulmonary embolism, and malignancies, as well as in fetal cell chimerisms in maternal serum and/or plasma. In this study, baseline concentrations of cell-free DNA in serum and plasma samples were evaluated for the study of posttransfusion chimerism. STUDY DESIGN AND METHODS: DNA was extracted from fresh or stored (4 degrees C for 1-6 days) normal donor serum or plasma samples (ACD; EDTA) by using reagents from an HIV assay kit. After incubation and washing of samples, purified DNA was amplified with HLA DQ-alpha primers (GH26 and 27) or human Y-chromosome primers (SA and SD) to quantitate the concentration of genomic DNA. RESULTS: Fresh serum samples had concentrations of cell-free DNA that were about 20-fold higher than the concentrations in fresh plasma samples. The concentration of cell-free genomic DNA in serum samples increased daily, to a level more than 100 times baseline after clotted blood tubes were stored at 4 degrees C for 4 to 5 days. There was a small increase in cell-free plasma DNA in stored ACD whole blood samples. Male WBCs, spiked into fresh nonanticoagulated female blood, were lysed during the process of clotting, with male DNA liberated into the serum samples. CONCLUSION: Most cell-free DNA in serum samples is generated during the process of clotting in the original collection tube. The concentration of cell-free genomic DNA in fresh plasma is probably the same as that in circulation. Consequently, while serum samples should not be used to monitor the concentration of cell-free DNA in a patient's circulation, serum collected from sample tubes containing clots (i.e., without anticoagulant), 3 to 5 days after the date of phlebotomy, could be useful as a source of DNA with which to screen for posttransfusion microchimerism.     

Maintenance of ATP Level of Incubated Human Red Cells by Controlling the pH

Fresh human blood was incubated at 37 C. in ACD solution, heparin-glucose solution, and heparin-glucose solution with p H controlled by automatic titration with sterile NaHCO₃ solution. In ACD solution glycolysis of the red cells slowed down with time and concomitantly the ATP level decreased sharply. In heparin-glucose solution, glycolysis continued more vigorously and the ATP levels were maintained longer. In the flasks in which the lactic acid was neutralized continuously, glycolysis proceeded almost unabated, and the ATP levels were correspondingly well maintained. Red cell viability and survival are presently assumed to be related to glycolytic maintenance of adequate levels of ATP. These studies demonstrate that storage of blood in heparin-glucose may prolong the red cell viability over that in ACD, and that neutralization of the lactic acid produced in glycolysis gives such dramatic improvement that a whole new system of blood storage may be predicated on this principle.     

A standardized technique for efficient platelet and leukocyte collection using the Model 30 Blood Processor

The Model 30 Blood Processor is a safe and simple means of harvesting blood cell components. Presently cell collection depends on a visual assessment by the operator of the indistinct boundaries of cell fractions. To determine when each cell component could best be harvested, serial samples were taken from the output port at fixed intervals anf the results of counts and differentials were graphed and tabulated. Studies in normal donors were done using acid-citrate- dextrose (ACD), 2 per cent sodium citrate in 6 per cent hydroxyethyl starch (HES), or heparin as anticoagulants. There was considerable overlap between the latter part of the platelet band, the leukocyte band and the rising hematocrit with all three anticoagulants. Normally functional lymphocytes could be harvested efficiently (approximately 80%) using ACD or heparin. Platelets could be harvested from ACD very efficiently (approximately 90%). Granulocytes could not be harvested from ACD (less than 10%) since they were dispersed in the red blood cell (RBC) layer. Using HES, granulocytes could be harvested efficiently (approximately 70%) by extending collection into the RBC layer. Based on these data, a standard technique for cell collection has been devised. The flow rate is slowed to 20 ml/min and collection is carried 30 ml (90 seconds at a rate of 20 ml/min) for platelets. The RBC loss is approximately 6 to 8 and 2 to 3 ml/pass respectively. These studies indicate that the Model 30 is a highly efficient apparatus for blood cell separation, but the volume of blood processed is limited by the intermittent blood flow.     

Effects of storage on in vitro platelet responses: Comparison of ACD and Na citrate anticoagulated samples

The present study was designed to evaluate whether the use of acid citrate dextrose (ACD) Formula A may enhance the survival of platelets during storage, thus allowing the continuance of platelet studies over the period of 2–3 hours usually recommended. For this purpose the effects of time on in vitro platelet response to several agonists have been investigated in platelet-rich plasma (PRP) obtained from blood samples anticoagulated with either Na citrate or ACD Formula A. The analysis of the data obtained in in vitro platelet aggregation studies using various parameters and at different time points demonstrated that storage of PRP obtained from citrated samples caused a marked reduction of platelet responses. This reduction was already evident after 6 hours, and a strong decrease was observed after 8 hours with all the agonists used. On the other hand, storage of ACD anticoagulated blood did not cause any significant decrease of platelet responsiveness up to 6 hours. A reduction of platelet aggregation became evident only after 8 hours, but not to the same extent as the one observed in citrated samples. Therefore, it may be concluded that the use of ACD Formula A as anticoagulant is capable of maintaining a normal platelet responsiveness up to 6–8 hours, thus permitting the investigation of platelet function for periods of time over those commonly recommended. © 1996 Wiley-Liss, Inc.     

Viability and function of outdated human red blood cells after biochemical modification to improve oxygen transport function, freezing, thawing, washing, postthaw storage at 4 C, perfusion in vitro through a bubble oxygenator, and autotransfusion

The quality of transfused blood is especially important during cardiac surgery, and red blood cell viability and function may be adversely affected during perfusion through the artificial blood oxygenator used during extracorporeal bypass. In this study, we administered 10 ml aliquot autotransfusions of rejuvenated red blood cells to 13 healthy volunteers after perfusion through an infant bubble oxygenator for one to three hours. Twenty-three other volunteers received rejuvenated red blood cells that had not been perfused. The red blood cells were biochemically modified after they had reached their outdating period, a process used to increase 2,3 DPG and ATP levels and improve oxygen transport function. The rejuvenated red blood cells were frozen with 40% W/V glycerol, stored frozen at -80 C for about 3 months, thawed, washed, and stored in a sodium chloride-glucose-phosphate solution at 4 C for as long as three days. Freeze-thaw recovery was about 97 per cent, and freeze-thaw-wash recovery about 90 per cent. Twenty-three units were transfused after 1 to 3 days of post-wash storage, and 13 units were perfused through an infant bubble oxygenator for as long as three hours before transfusion. The 24-hour posttransfusion survival values were about 80 per cent and oxygen transport function was either normal or improved whether or not the units were perfused before transfusion.     

Damage to Red Cells Due to Their Collection and Storage in ACD

Experiments were performed using aliquots of the same blood incubated at 37 C. in spinner flasks under 95%N₂ – 5% CO₂. Various anticoagulant systems and p H's were compared, using the rate of glycolysis and ATP levels as criteria of metabolic activity. Citrate and lower p H's were detrimental to glycolysis and the maintenance of red cell ATP levels. Non-citrate systems that were closer to normal blood p H were superior to ACD, by the criteria used in this in vitro study. The addition of phosphate, as in the citrate-phosphate-dextrose system, appeared beneficial.     

Acid citrate dextrose (acd) formula a as a new anticoagulant in the measurement of in vitro platelet aggregation

To evaluate whether the use of ACD Formula A may affect in vitro platelet function, blood samples were obtained from 21 healthy blood donors and anticoagulated in ACD (acid-citrate dextrose, NIH Formula A), Na citrate 3.8%, and K₃EDTA. Platelet count, mean platelet volume, and in vitro platelet aggregation were evaluated on each sample. No significant difference was observed in platelet count and mean platelet volume among the different samples. Conversely, the ACD treated platelets showed a higher reactivity to the agonists as demonstrated by a significant increase of the maximum percentages of aggregation induced by ADP, epinephrine, and collagen, as well as a significant decrease of secondary aggregation thresholds to ADP and epinephrine. In conclusion, it may be speculated that ACD Formula A is capable of better maintaining the intraplatelet signal transduction mechanisms during PRP preparation, thus improving the overall responsiveness of platelets.©1995 wiley-Liss, inc.     

Studies on Bank Blood Collected and Stored under Various Conditions with Particular Reference to Its Use in Open Heart Surgery

A few of the factors responsible for the preservation of the characteristics of fresh blood during storage have been studied, with particular reference to the provision of donor blood for open heart surgery. It has been indicated that present data leave us no reason to think that platelet viability and the level of labile clotting factors in stored blood will be any different whether the blood is collected by gravity or with the aid of vacuum. Changes in bloods stored at 4C. for a 21 day period were investigated with ACD, heparin and Edglugate-Mg as the anticoagulants. The studies on heparinized blood suggest that after two or three days, the fall in platelets and the rise in plasma potassium and hemoglobin are so much greater than occur in ACD that storage for longer periods than that will probably never be safe. On the other hand, in vivo studies indicate that 24-hour-old heparinized blood is just as safe to use as four-hour-old blood. Edglugate-Mg was developed in an effort to get around the limited storage period of heparinized blood. Results reported here show that plasma potassium levels rise very rapidly in this type of blood. Platelets are not well-preserved in vitro , and Factor V levels are apparently low soon after collection.     

Comparison Studies of Whole Blood Stored in ACD and CPD and with Adenine

Using 70% 24-hour posttransfusion survival as one of the criteria of preservation, ACD and CPD anticoagulant solutions with and without adenine were tested after storage for 28, 35, or 42 days. At 28 days, all solutions had average survivals of 70% or better. The average survival values for the ACD and CPD solution groups were less than 70% at 35 and 42 days. However, both solutions with adenine had a group average value over 70% even after 35 or 42 days of storage. The average survival values between the two anticoagulants alone, were not significantly different at each time period. The two anticoagulants when adenine was included had increased survival and there was no statistical difference between the levels. In comparing two units obtained from the same subject, survival percentages were significantly higher in almost every recipient when the adenine-supplemented stored blood was used. Other chemical determinations did not show significant alterations and no toxic effects were observed in the recipients.
Since all units containing adenine had survival values greater than 70% in the 28- and 35-day periods, these units would appear to have been effectively preserved and blood stored under these conditions could be used in routine transfusions, reserving units stored 42 days for emergency use.     

Calcium Ion Activity During Rapid Exchange Transfusion with Citrated Blood

Measurement of ionized calcium levels by the technic of Soulier during massive transfusion of ACD blood supplemented with various quantities of calcium led to the apparently erroneous conclusion that ionized calcium levels would rise in the recipient when each unit was supplemented with 0.6 g CaCl₂, the quantity which restores normal ionized calcium levels in vitro .
Measurement of ionized calcium activity with the Orion electrode indicates that this ratio of CaCl₂ will maintain an approximately normal ionized calcium level in dog recipients if it is infused simultaneously with the blood. Based on this information, it appears reasonable to assume that heparinized ACD blood, recalcified with 0.6 g CaCl₂ per unit, may be used to prime the extracorporeal circuit for open heart surgery with the confidence that it will not alter the ionized calcium activity of the patient's blood.
Addition of calcium to the circulation following administration of ACD blood causes a sharp rise and then a fall in the recipient's level of ionized calcium. Further studies in a variety of situations are required before final guidelines can be formulated for supplementation of ACD blood with calcium in massive transfusion.     

Accumulation of DI-2-Ethylhexyl Phthalate (DEHP) in Whole Blood, Platelet Concentrates, and Platelet-Poor Plasma

The concentration of the plasticizer, di-2-ethylhexyl phthalate (DEHP), in the plasma was measured after storage of the whole blood in polyvinylchloride plastic bags at 4 C for up to 38 days in either ACD or CPD. The plasma from ACD-stored whole blood contained more DEHP than that from CPD-stored whole blood. The continuous-flow centrifugation washing procedures removed about 98 per cent of the DEHP from ACD whole blood stored for 33 days at 4 C.
DEHP was assayed in the platelet-rich plasma, platelet-poor plasma, supernatant of the platelet concentrate, washed platelets, and washed red blood cells prepared from CPD whole blood. Very little DEHP was found in the washed red blood cells and platelets, a small amount was found in the platelet-poor plasma, and a large amount in the supernatant of the platelet concentrate. A greater amount of DEHP accumulated in the platelet concentrates that were stored at 22 C than in those stored at 4 C. When platelet concentrates from CPD whole blood were stored at 22 C for 72 hours, the amount of DEHP was about four times that observed after 4 C storage for the same length of time.