Effect of citrate anticoagulants on factor VIII levels in plasma

The citrate anticoagulants used during blood collection have been developed for their benefits to red cells. The concentrations in which they are used are strictly regulated in the United States: citrate-phosphate-dextrose-adenine (CPDA) is used in a 1:8 ratio for the collection of whole blood, whereas 4 percent sodium citrate (NaCit) is used in a 1:10 ratio for manual plasmapheresis. Acid-citrate-dextrose formula A (ACD-A) or formula B (ACD-B) and NaCit are commonly used in a 1:12 or 1:15 ratio during automated plasmapheresis. These anticoagulants have different initial and final pH values and citrate concentrations and different effects on the recovery of factor VIII (FVIII) in the plasma. NaCit has a higher initial pH (6.64) than ACD-A (4.98), ACD-B (5.60), or CPDA (5.12). The effects of these different anticoagulants on plasma constituents obtained from six healthy subjects were studied. In standard citrate concentrations, the FVIII level was significantly lower (p less than 0.05) in the NaCit used for manual plasmapheresis than in either of the ACD solutions used in automated plasmapheresis (104 U/dl vs. 153 and 160 U/dl). When various ratios of NaCit to blood were used, the pH increased from 7.62 at a 1:10 dilution to 7.65 at a 1:50 dilution. As expected, a progressive decrease in anticoagulant level was associated with an increase in ionized calcium and also in the level of FVIII, with the latter values rising from 104 U per dl at 1:10 to 137 at 1:20 and 148 U per dl at 1:30. Clot formation was detected only at a ratio of 1:35.(ABSTRACT TRUNCATED AT 250 WORDS)     

Use of hydroxyethyl starch to improve granulocyte collection in the Latham blood processor

This report describes a practical, relatively inexpensive system of leukapheresis and plateletpheresis, utilizing the Haemonetics blood processor. The results of 236 phereses of normal donors are reported, with particular attention to an evaluation of five different anticoagulant mixtures: ACD, 2 per cent citrate in saline, and three different combinations of citrate and hydroxyethyl starch (HES). These mixtures were compared with respect to their effectiveness in the harvesting of granylocytes and platelets, respectively. The best harvest of granulocytes (3.5 × 10(9) cells per liter of blood processed) was with 6 per cent HES anticoagulated with trisodium citrate. All solutions gave about the same platelet yields (a mean of 1.6 × 10(11) per liter of blood processed). Because of its higher citrate content, ACD caused three times as many donor reactions as the other solutions. The use of HES thus permits this widely available plateletpheresis system to be used for leukapheresis with only minimal procedural modification.     

The Anticoagulants of Choice for Platelet Transfusions

A study was made of the recovery of Cr⁵¹-labeled platelets transfused to normal recipients using various anticoagulant solutions. With ethylenediamine-tetra-acetate (EDTA) recoveries averaged 27 per cent (range 15% to 35%). With ACD (N.I.H. formula A) recoveries averaged 37 per cent (range 34% to 41%). With a citrate solution containing additional acid so as to reduce the p H of platelet-rich plasma to 6.5 before concentration of cells, recoveries averaged 62 per cent (range 37% to 81%). Platelet clumping, usually a problem when platelets are concentrated in citrate, was prevented at p H 6.5. It is suggested that a citrate solution containing sufficient acid to buffer platelet-rich plasma at p H 6.5 will increase the effectiveness of platelet concentrates.     

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.     

A Clinical Evaluation of the Use of Citrate-Phosphate-Dextrose Solution in Children

Red cells collected in CPD anticoagulant have been shown to have a mean postinfusion survival of greater than 75 per cent after 28 days of storage at 4C, in contrast to blood collected in ACD solution whose mean survival is less than 70 per cent after 28 days.
In a clinical study, 3,704 units of CPD blood collected in double plastic blood packs were utilized for 4,354 transfusions in 2,425 children. Evaluation of the stability of red cell antigens disclosed that blood preserved in CPD showed a minimal loss of antigenicity after 28 days. In contrast, clotted samples were hemolyzed and demonstrated marked loss of antigenicity for Kell, hr'(c) and P after 14 days.
Whole blood preserved in CPD provides two basic advantages over the presently used ACD solution. It provides a more physiologic blood for the recipient and has decreased the outdating percentage by better than 65 per cent.     

Comparative Viability of Blood Stored in ACD and CPD

The normal distribution of red blood cell viability for ACD and CPD units stored for 21 days was studied. For ACD and CPD units, respectively (n=41,37), the means and standard deviations were as follows: 24-hour survival, 75.7 + 6.2 and 79.4 + 6.4; early recovery, 91.6 + 3.6 and 94.1 + 3.5; t/2, 29.4 + 3.0 and 27.9 + 4.3. Early recovery and survival were significantly higher for CPD, but more important than the difference in mean survival is that by current standards of acceptability, the incidence of donors who will be deemed undesirable is approximately 6 per cent for CPD, as opposed to 20 per cent for ACD. Neither early recovery, 24-hour survival, nor t/2 could be shown to correlate with pH, plasma potassium, plasma sodium, per cent hemolysis, and osmotic fragility. The mean and standard deviation of survival for 18 units of 28-day-old CPD blood was 70.7 + 11. Since the standard error was large, the frequency distribution could not be determined, and the number of units with survivals that would fall below the minimum standard could not be ascertained. Nevertheless, comparison with 21-day old ACD did not show a significant difference in the mean survival, although the range observed was much wider. The results also point out the need for greater number of observations with increasing duration of storage for adequate appraisal of blood preservative solutions.     

The effect of citrate anticoagulants on apheresed plasma

Platelet-rich plasma (PRP) was apheresed from the same donors into four anticoagulant formulations favoured for preparing platelet concentrates (PC), and platelet-poor plasma (PPP) harvested by a second centrifugation. A fifth control collection was made by direct apheresis of PPP. Matched 3–4 kg pools of PPP underwent partial fractionation to determine the effects of plasma anticoagulant on factor VIII recovery and the potential thrombogenicity of prothrombin complex Although an excellent factor VIII recovery was achieved in each case, the cryoprecipitate from plasma apheresed into acid citrate dextrose formula B (ACD-B) showed evidence of incipient clotting, and the factor IX concentrate from the same plasma contained a high concentration of activated factors. It was concluded that ACD-B, in the ratio used, provides insufficient citrate for secure anticoagulation and should not be used when PPP is destined for fractionation of coagulation factor concentrates.     

Human blood components. I. Preparation and characteristics of leukocyte concentrates from single units of human blood

The distribution of white blood cells in the packed cells remaining after removal of platelet-rich plasma has been examined. The top 50 ml, which includes the huffy coat, was expressed in 5 aliquots of 10 ml and analyzed. With ACD, the top SO ml contained 52 per cent of the total sedimented WBC but only 33 per cent of the available polymorphonuclear leukocytes (PMN). With CPD, the top 50 ml contained 65 per cent of the total sedimented WBC and 52 per cent of the available PMN. The PMN were relatively evenly distributed throughout the five aliquots, while the non-PMN leukocytes were concentrated in the first three aliquots. The hematocrit of the top layer of packed cells was consistently lower for units collected in ACD than for units collected in CPD anticoagulant. Standard curves for the volume dependence of the differential and hematocrit were generated from these data. The characteristics of leukocyte concentrates routinely prepared as part of component production from the top 35 ml of the packed cells have been analyzed. Leukocyte concentrates prepared with ACD had, on the average, lower hematocrit, a lower percentage of PMN and fewer total white blood cells than leukocyte concentrates prepared from CPD-anticoagu-lant. Using CPD-anticoagulant instead of ACD-anti-coagulant during source blood collection resulted in an increase of the mean PMN differential count of the leukocyte concentrates from 29.8 to 45.7 per cent and of the mean total number of PMN in the concentrates from 0.46 × 10⁹ to 0.74 × 10⁹. The application of these data to component preparation and use is discussed.     

Adenine in Blood Preservation: Posttransfusion Viability and Biochemical Changes

Posttransfusion viability was studied in red blood cells stored for 21 days in ACD solution and for 35 days in ACD solution supplemented with adenine to a final concentration of 0.5 mM. The survival of radio-chromium-labeled red cells was determined after transfusion of 10 ml. of autologous blood and 350–400 ml. homologous blood. The viability values were about the same for the two transfusion procedures. The mean posttransfusion viability was 80 per cent for erythrocytes stored for 35 days in the medium containing adenine and 79 per cent for cells preserved in ACD solution for 21 days.
The concentration in the erythrocytes of ATP, ADP, AMP, reduced and oxidized glutathione, 2,3-diphosphoglycerate, and four glycolytic enzymes was measured before and after storage in the two media. The ATP and the total adenine nucleotide concentrations were much higher in the red cells stored in the adenine-containing solution. Of the enzymes tested, only phosphofructokinase decreased in activity during 35 days of storage. The decrease was about 50 per cent and was not dependent on the storage solution.
This study supports the theory that decreased adenine supply is an important cause of damage to erythrocytes in ACD solution.     

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.     

Observations on Recipient Plasma Hemoglobin Concentration after Transfusion with Glycerolized Frozen Blood

Recipient plasma hemoglobin was measured prior to, during, and following infusions of deglycer-olized red blood cells resuspended in autologous plasma, 5 per cent outdated albumin resuspension medium, and isotonic saline. Blood collected in ACD solution and stored at 4 C was studied in a similar manner. The effect of aggregation and disaggregation of erythrocytes on the recipient plasma hemoglobin was observed. Observations in 12 nonsurgical recipients indicate that the mode of removal of nonviable deglycerolized erythrocytes is primarily through an extravascular mechanism without detectable hemoglobinemia.
The aggregation of erythrocytes in either ACD blood or deglycerolized, reconstituted blood with 5 per cent dextrose and water and their disaggregation with 0.9 per cent sodium chloride in a common channel of intravenous tubing produced no unexplained elevation of the recipient plasma hemoglobin. In seven recipients studied, no abnormal antibodies were detected following repeated transfusions of deglycerolized erythrocytes over a period of time.     

Studies on Stored Whole Blood II. Use of Packed Red Blood Cells

Whole blood collected in ACD or ACD with adenine was studied as units stored with or without plasma over 14 to 28 days. Comparisons were made using various tests, with posttransfusion survival in vivo being the primary guideline. The adverse effects which resulted from storage appeared more slowly in units with plasma than in units without plasma. The inclusion of adenine to either packed or nonpacked units was associated with even less evidence of storage damage. Using the criterion of over 70 per cent recovery in 24 hours as signifying adequate preservation, the average survival value for each group was better when the plasma was present, and even higher when adenine had been added. All tested groups had average values exceeding 70 per cent recovery after 21 days of storage. Blood stored for 28 days with adenine had over 70 per cent recovery with or without plasma. Though the storage damage became more apparent in units without plasma, the average survival value would suggest that the packed units were still useful after 21 days of storage, with extension to 28 days when ACD with adenine was used.     

A comparison of factor VIII activity in cryoprecipitates prepared from ACD and CPD plasma

In an attempt to develop a method to produce a cryoprecipitate with a predictable Factor VIII potency, several variables were studied and statistically analyzed. These included the hematocrit, age, blood group and Rh type of the donor; possible epinephrine release; the degree of lipemia, volume, pH and Factor VIII activity of the donor plasma; the volume of cryoprecipitates, its Factor VIII activity and the per cent yield; and the Factor VIII activity of the supernatant plasma. Cryoprecipitates were prepared by the method of Pool and Shannon from the plasmas of 40 random male donors, half the bloods being drawn in ACD and half in CPD. None of the variables had a significant influence on the per cent yield of activity, although the mean values obtained suggest that the per cent yield of Factor VIII activity in the cryoprecipitates prepared from CPD plasmas is higher than in those prepared from ACD plasma. Although the per cent yield of Factor VIII in cryoprecipitates prepared from CPD plasma is not significantly different from that of ACD plasma, the mean total units of Factor VIII activity is significantly higher in CPD cryoprecipitates. It also was confirmed that a higher per cent Factor VIII activity in the donor results in a relatively higher activity in the cryoprecipitate. The data indicate that if CPD plasma collected from donors having above a certain minimum per cent activity were used for cryoprecipitate production, one could be assured of having a minimum of 100 units of Factor VIII activity per bag.     

The use of calcium chelating agents and prostaglandin E to eliminate platelet and white blood cell losses resulting from hemoperfusion through uncoated charcoal, albumin-agarose gel, and neutral and cation exchange resins.

Hemoperfusion through adsorbents such as charcoal, cation exchange (e.g., AG 50W-X8) and uncharged (e.g., XAD-2) resins, and albumin-agarose gel (AAG) has been proposed for use in patients with hepatic failure. However, the loss of white blood cells and, particularly, platelets caused by each of these adsorbents remains a major deterrent to their clinical use. In vitro studies demonstrate that addition of citrate, ethylenediamine tetraacetate (EDTA), or oxalate to heparinized human blood eliminated this loss of formed blood elements during hemoperfusion. The improvement in postperfusion platelet counts (per cent of preperfusion values) produced by citrate were XAD-2, 13 leads to 95 per cent; AG 50W-X8, 10 leads to 94 per cent; AAG, 17 leads to 94 per cent; and charcoal, 44 leads to 96 per cent. Prostaglandin E1 in high doses (5 mug/ml.) markedly reduced platelet losses. Lower doses were less uniformly effective. Three young rhesus monkeys were hemoperfused for 160 minutes with columns containing AAG, XAD-2, and charcoal. During the first 80 minutes, citrate and calcium were infused into the column inflow and outflow lines respectively. For all three adsorbents, average platelet counts in the monkeys (115 per cent) and column effluent (95 per cent) were unchanged from preperfusion control values during the first 80 minutes but fell promptly to 13 and 7 per cent, respectively, after the citrate infusion was stopped. Each of the monkeys tolerated the procedure without ill effects. Use of a system analogous to that described here may facilitate clinical application of the technique of hemoperfusion through a variety of adsorbents.     

The in vivo survival of red blood cells stored in modified CPD with adenine: report of a multi-institutional cooperative effort

In order to provide data in support of licensure applications for citrate-phosphate-dextrose (CPD) supplemented with adenine, a multi- institutional cooperative effort was organized to determine survivability of red blood cells subjected to prolonged liquid storage. Two manufacturers supplied plastic multiple bag blood storage containers prefilled with modified CPD (glucose 25/ greater than the normal concentration) supplemented with adenine (17.0 to 17.3 mg per 63 ml of anticoagulant; 0.25 millimolar approximate final concentration when diluted with 450 ml of whole blood for 35 days showed a mean survival of 80.53 +/- 6.44 per cent (1 SD). Both red blood cell and supernatant plasma biochemical characteristics were comparable to those reported for whole blood stored for 21 days in either acid-citrate- dextrose (ACD) or CPD. Red blood cells from 19 units stored as concentrates for 35 days (Hct 75.03 +/− 3.74%) had a mean survival of 71.38 +/− 10.3 per cent with considerable interdonor variation in survival and interlaboratory variation in some biochemical characteristics. Red blood cells from eight units stored as concentrates (Hct 75.38 +/− 4.30%) for 28 days showed a mean survival of 83.97 +/− 6.10 per cent and biochemical characteristics comparable to those reported for red blood cell concentrates stored in CPD or ACD for 21 days. Modified CPD with adenine as formulated offers an improved anticoagulant for blood banking by extending the permissible red blood cell storage period.     

Citrate-Phosphate-Dextrose Solution for Preservation of Human Blood: A Further Report

A comparison of clinically significant in vitro characteristics of ACD and CPD blood is presented.
In clinical trials conducted at three hospitals, 586 units of blood collected in CPD in plastic bags were utilized for 622 individual treatments as single units or subdivided aliquots of whole blood, and as packed red cells in 332 patients. The expiration period was 28 days, and five bloods were discarded as outdated. There was one reported pyrexial reaction and one proven bacterial contamination.
Forty-two per cent of the bloods or components were issued during the first week, 43 per cent during the second and third weeks, and 15 per cent during the fourth week of storage.
Of the total bloods or components issued, 9 per cent of the whole bloods, 25 per cent of the subdivided blood aliquots, and 23 per cent of the packed red cells were transfused during the fourth week of storage.
No difficulty with clotting was observed during collection, storage, or transfusion.     

Clinical Evaluation of Transfused Blood after Long-term Storage in ACD with Adenine

Whole blood collected in ACD with or without adenine was transfused into patients with various forms of blood loss. The units collected in ACD alone had been stored 14 to 28 days, while those units collected in ACD with adenine had been stored 28 to 42 days. Of the 771 units in the study, 545 were transfused into 295 patients. A total of 276 units were collected in ACD with adenine and were given to 130 patients. Little or no reaction was detected from clinical observation and selected laboratory tests related to the transfusion and/or solution. Isotope labeling of some of the transfused units given to more than 80 patients provided an estimate of the 24-hour posttransfusion survival and the results were similar to the average survival obtained in normal subjects after single-unit transfusions. Based on clinical laboratory findings, apparent lack of toxicity combined with adequate support of the bleeding patient suggest that blood stored in adenine is useful after longer storage periods than is blood stored in plain ACD. The 70 per cent 24-hour survival guideline suggests satisfactory survival after 35 days of storage, and possibly after 42 days.     

Specific Agglutinability of Erythrocytes from Whole Blood Stored at 4 C

When whole blood is supplemented with adenine, the erythrocytes are suitable for transfusion after at least 35 days of storage at 4C, but information is not available as to whether such cells remain satisfactory for blood typing and pre transfusion tests for evidence of incompatibility. Accordingly, we designed experiments to evaluate by AutoAnalyzer the specific agglutinability of erythrocytes obtained from whole blood stored at 4C. Seven normal volunteer donors of both sexes, aged between 22 and 31 years, were chosen and bled periodically so that, over a three-day testing period, all blood specimens from each donor could be evaluated. Each donor provided at each bleeding sufficient blood to permit aliquots of 20 ml to be stored as clotted blood and 8 ml to be stored as citrated whole blood. Four kinds of citrate solution were used: ACD Formula A, CPD, ACD supplemented with adenine, and CPD supplemented with adenine. All clotted specimens were tested after 0, 1, 2, 3, 4, and 5 weeks of storage, while all citrated specimens were tested after 0, 1, 2, 3, 4, 5, 6, 8, and 10 weeks of storage. Six blood group systems were used for evaluation with the following reagents: anti-A or anti-B, anti-Rhl (Rh_o or D), anti-K2 (k or Cellano), anti-Jk^a, anti-Fy^a or anti-Fy^b, and anti-M. Each reagent was used at a dilution to support from 20 to 80 per cent agglutination with freshly drawn blood, and each reagent was tested under two conditions: low ionic and normal ionic. The results disclosed loss of specific agglutinability in association with the time of storage of whole blood at 4 C. The onset and degree of loss depended both on the kind of test used (normal ionic tests showed loss 2.8 weeks earlier than did low ionic tests) and the condition in which the blood was stored (loss occurred sooner and then progressed more with clotted blood than with citrated blood). Average losses of 25, 50, and 75 per cent were observed at 1.1, 2.1, and 4.0 weeks of storage for normal ionic tests of clotted blood, and at 4.1, 6.0, and 7.6 weeks for similar tests of citrated blood. Loss of specific agglutinability during storage was significantly less for the red blood cells of some donors than for others, but systematic differences between specific blood-typing tests were not observed. ACD and CPD solutions behaved similarly, while adenine supplementation exerted a slight preservative effect after 6, 8, and 10 weeks of storage. Loss of specific hemagglutinability of stored cells was not due to loss of specific antigenic sites because stored cells were as effective as fresh cells for the absorption of anti-B, anti-Rhl, and anti-M. Loss might, however, be related to an increase in the accumulation of IgG, fibrinogen, and albumin at the surfaces of erythrocytes.     

The impact of citrate concentration on adhesion of platelets and leukocytes to adsorbents in whole blood lipoprotein apheresis

Lipoprotein apheresis is applied to deplete low density lipoprotein and other apolipoprotein B containing lipoproteins in patients with severe familial hypercholesterolemia, hypertriglyceridemia associated pancreatitis, or lipoprotein (a)‐hyperlipoproteinemia. Anticoagulation of the extracorporeal circuit may influence cellular activation, as evidenced by a reduction of inflammatory parameters during regional citrate anticoagulation with acid citrate dextrose A (ACD‐A) commonly used in whole blood lipid apheresis. While the citrate concentration in the extracorporeal circuit has to ensure efficient anticoagulation, citrate infusion into the patient should be limited to avoid citrate overload. We assessed the influence of citrate concentration on cellular activation during in vitro circulation of whole blood containing 2.8 mM citrate (ACD‐A 1:40), 5.6 mM citrate (ACD‐A 1:20), or 13 mM citrate over polyacrylate‐based adsorbents for lipoprotein apheresis. We found increased platelet adhesion for anticoagulation with 2.8 mM citrate as compared to 5.6 or 13 mM citrate, as shown by cell counting and confirmed by scanning electron microscopy of adsorbent beads as well as by elevated levels of platelet activation markers and of platelet‐derived microvesicles. Leukocytes showed an equivalent adhesion pattern, while red blood cells remained unaffected at all citrate concentrations. Passage of blood over two consecutive columns resulted in enhanced platelet adhesion to the second column, presumably due to upstream preactivation. In conclusion, citrate influences activation and adhesion of platelets and leukocytes in a concentration‐dependent manner, and ACD‐A 1:20, equivalent to a citrate concentration of 5.6 mM in whole blood, ensures minimal cellular activation during passage of whole blood over polyacrylate‐based adsorbents.     

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.