Warm storage of whole blood for 72 hours

BACKGROUND: In field emergency medicine, fresh whole-blood units are stored at room temperature up to 24 hours or occasionally longer. Few data exist on the integrity and in vitro functional properties of whole blood stored warm beyond 24 hours. STUDY DESIGN AND METHODS: Ten citrate phosphate dextrose solution whole-blood units were collected and divided into two equal volumes. One-half of each unit was stored at 19 degrees C and the other half was stored at 25 degrees C, encompassing the accepted range for room temperature storage. At 6, 24, 48, and 72 hours, aliquots were collected from each unit and whole blood analyzed for cell counts, gases, and clotting function with thromboelastography, red cells for intracellular analytes, platelet (PLT)-rich plasma for aggregometry, and the supernatant for hemoglobin, potassium, glucose, lactate, and plasma clotting studies. RESULTS: Whole-blood units stored at room temperature maintained cellular counts and coagulation activity for up to 72 hours. Units stored at 19 degrees C demonstrated greater RBC adenosine triphosphate and 2,3-diphosphoglycerate (DPG) content and stronger responses in PLT aggregation studies when compared with 25 degrees C storage. No significant hemolysis was observed, and no bacterial growth was detected. CONCLUSION: Storage of whole blood at room temperature for 72 hours leads to marked reductions in pH and DPG, but the observed reduction in PLT function and plasma coagulation factor activity was surprisingly modest compared to literature values. These findings should prompt additional investigation, given their potential importance for whole blood processing and field-expedient transfusion.     

Pyrophosphate as a novel anticoagulant for storage of whole blood: A proof-of-concept study

Background

Citrate is the only anticoagulant currently Food and Drug Administration (FDA)-approved for the long-term storage of blood for transfusion. Citrate inhibits phosphofructokinase and may play a pro-inflammatory role, suggesting that there may be an advantage to using alternative anticoagulants. Here, we examine the use of pyrophosphate as an anticoagulant.

Study design and methods

Whole blood samples from healthy donors were anticoagulated either with citrate–phosphate–adenine–dextrose (CPDA-1) or our novel anticoagulant mixture pyrophosphate-phosphate–adenine–dextrose (PPDA-1). Samples were assessed for coagulation capacity by thromboelastography immediately after anticoagulation (T0) with and without recalcification, as well as 5 hours after anticoagulation (T1) with recalcification. Complete blood counts were taken at both timepoints. Flow cytometry to evaluate platelet activation as well as blood smears to evaluate cellular morphology were performed at T1.

Results

No clotting was detected in samples anticoagulated with either solution without recalcification. After recalcification, clotting function was restored in both groups. R-Time in recalcified PPDA-1 samples was shorter than in CPDA-1 samples. A reduction in platelet count at T1 compared to T0 was observed in both groups. No significant platelet activation was observed in either group at T1. Blood smear indicated platelet clumping in PPDA-1.

Conclusion

We have shown initial proof of concept that pyrophosphate functions as an anticoagulant at the dose used in this study, though there is an associated loss of platelets over time that may limit its usefulness for blood storage. Further dose optimization of pyrophosphate may limit or reduce the loss of platelets.     

两种从陈旧血中抽提基因组DNA方法的比较

目的比较盐析法和磁珠法从陈旧血中抽提基因组DNA的效果和特点,以便常规用于骨髓库HLA基因分型。方法使用TECAN全自动工作站为平台,分别用Gentra盐析法DNA抽提试剂盒和Promega磁珠法DNA抽提试剂,从48份陈旧全血样本中抽提基因组DNA,提取的基因组DNA均溶于100μlTE中;然后用紫外分光光度计测定其产量和纯度,并用琼脂糖凝胶电泳法测定DNA样本的完整性。结果从400μl陈旧全血中提取基因组DNA,用盐析法获得的DNA含量为28.6±8.7ng/μl,A260/A280值平均为1.63±0.209;用磁珠法获得的DNA含量为94.2±10.3ng/μl,A260-A320/A280-A320值平均为1.821±0.102;琼脂糖电泳法检测表明两种方法提取的DNA的分子量均大于15kb。结论使用磁珠法从陈旧血中所获得的DNA的产量和纯度明显高于盐析法,适合于骨髓资料库陈旧血样本的常规HLA基因分型实验以及下游的分子生物学实验。     

两种从陈旧血中抽提基因组DNA方法的比较

目的 比较盐析法和磁珠法从陈旧血中抽提基因组DNA的效果和特点,以便常规用于骨髓库HLA基因分型.方法 使用TECAN全自动工作站为平台,分别用Gentra盐析法DNA抽提试剂盒和Promega磁珠法DNA抽提试剂,从48份陈旧全血样本中抽提基因组DNA,提取的基因组DNA均溶于100 μlTE中;然后用紫外分光光度计测定其产量和纯度,并用琼脂糖凝胶电泳法测定DNA样本的完整性.结果 从400 μl陈旧全血中提取基因组DNA,用盐析法获得的DNA含量为28.6±8.7 ng/μl,A260/A280值平均为1.63±0.209;用磁珠法获得的DNA含量为94.2±10.3 ng/μl,A260-A320/A280-A320值平均为1.821±0.102;琼脂糖电泳法检测表明两种方法提取的DNA的分子量均大于15 kb.结论 使用磁珠法从陈旧血中所获得的DNA的产量和纯度明显高于盐析法,适合于骨髓资料库陈旧血样本的常规HLA基因分型实验以及下游的分子生物学实验.     

Active cooling of whole blood to room temperature improves blood component quality

BACKGROUND: Many countries use cooling plates to actively cool collected whole blood (WB) to room temperature. Until now, no paired comparison had been performed, and it was our aim to compare the effect of active versus no active cooling on the in vitro quality of WB and subsequently prepared blood components. STUDY DESIGN AND METHODS: Two units of WB were pooled and divided shortly after donation. One unit was placed under a butane‐1,4‐diol plate to obtain active cooling; the other was placed in an insulated box with other warm units to mimic worst‐case holding conditions. WB was held overnight and processed into a white blood cell (WBC)‐reduced red blood cells (RBCs), buffy coat (BC), and plasma. The BCs were further processed into platelet (PLT) concentrates. RBCs were stored for 42 days, and PLT concentrates for 8 days (n = 12 paired experiments). RESULTS: After overnight storage, ATP content of the RBCs was 4.9 ± 0.3 µmol/g Hb for actively cooled WB versus 4.5 ± 0.4 µmol/g Hb for not actively cooled WB (p < 0.001). On Day 42 of storage, RBCs prepared from this WB contained 3.1 ± 0.3 µmol ATP/g Hb with active cooling versus 2.6 ± 0.3 µmol/g Hb without (p < 0.001). Hemolysis on Day 42 was 0.35 ± 0.08% with active cooling and 0.67 ± 0.21% without (p < 0.001). No effect was observed on the in vitro quality of plasma, BC, or PLT concentrates. CONCLUSIONS: Active cooling of WB results in improved ATP levels and less hemolysis in WBC‐reduced RBCs, although the clinical implications are unclear. It has no effect on the in vitro quality of plasma or PLT concentrates.     

Simultaneous detection of whole blood chemiluminescence in microtitre plates.

The measurement of reactive oxygen species provides a simple method for monitoring the degree of activation of leukocytes in various disorders, and for determining the effects of drugs on this activation. The present report describes the determination of luminol- or lucigenin-amplified chemiluminescence of whole blood in a microtitre plate assay with a 96-well luminometer (HAMAMATSU MTP reader). Using heparinized venous human blood from healthy donors, optimal chemiluminescence intensities were determined at a blood dilution of 1/100 in a total volume of 0.25 ml of Hank's balanced salt solution, containing 0.4 mmol/l luminol as enhancer and either opsonized zymosan (1 milligram) or the phorbol ester, 12-O-tetradecanoylphorbol 13-acetate (10(-6) mol/l), as stimuli. The in vitro effects of nordihydroguaiaretic acid, diphenylene iodonium, and diclofenac were tested. After preincubation of the diluted whole blood with these drugs for 15 min, the zymosan-stimulated chemiluminescence was diminished in all cases. The specific NADPH oxidase inhibitor, diphenylene iodonium, was most effective (half maximal inhibition at 1.5 x 10(-8) mol/l), whereas higher concentrations of the antioxidant, nordihydroguaiaretic acid (1.6 x 10(-6) mol/l), or the nonsteroidal antiinflammatory drug, diclofenac (about 10(-5) mol/l), were needed to achieve half maximal inhibition. In addition to its usefulness in the rapid screening of drug effects this assay system seems to be very beneficial for the clinical diagnosis of congenital disorders. Furthermore, it is suited as an effective and simple method for the continuous determination of the phagocyte functional state in patients in pathophysiological situations and during therapy.     

Infusion flow rates of whole blood and AS-1-preserved erythrocytes: a comparison

We compared the flow rates of whole blood and erythrocytes resuspended in a new preservative solution (AS-1, consisting of adenine, dextrose, mannitol, and saline) which results in an erythrocyte preparation with a hematocrit lower than that of packed erythrocytes. When 100 ml of AS-1 solution is added to erythrocytes, a hematocrit of 59 +/- 5% is consistently obtained, and the resultant product has an improved flow rate. When we compared the infusion flow rates of whole blood and AS-1-preserved erythrocytes in vitro and in vivo, we found that flow times were shorter for AS-1 erythrocytes than for whole blood in vitro and in vivo, the flow rates of AS-1 erythrocytes and whole blood when expressed per volume were similar in vivo, and the flow rate of AS-1 erythrocytes for erythrocyte mass delivery in vivo was superior to that of whole blood. Thus, we conclude that the flow rates of the two products are comparable.     

In vitro comparison of CPD whole blood with conventional blood components

IntroductionResuscitation of severely injured trauma patients is commonly performed using red blood cells in additive solution supplemented with plasma and platelet concentrates. There is an increasing interest in the use of low anti-A titer Group O whole blood (LTOWB) in the early management of the resuscitation. It is unclear whether clinical outcome is improved using this approach.MethodsExpired units of CPD-LTOWB were studied on Day 22 and expired units of thawed plasma on Day 6 and Day 7. LTOWB was assessed for hemoglobin content, clotting factor levels and platelet numbers and function using thromboelastography (TEG) and impedance aggregation. Assays of fibrinogen and FV, FVIII, FVII and FX were performed on the expired plasma. The LTOWB hemoglobin was compared to red cells in additive solution (AS-RBCs) and the clotting factor levels to those of expired thawed plasma. Platelet function was compared to fresh whole blood samples from healthy subjects.ResultsLTOWB contained slightly more hemoglobin than the AS-RBCs (Medians, 66 v 59 G), and the plasma content of fibrinogen was similar. Other clotting factors were reduced by approximately 15% except for FVIII which was 30% less. Both TEG and impedance aggregometry showed evidence of residual platelet function despite the prolonged period of refrigerator storage.ConclusionLTOWB contains higher hemoglobin and adequate clotting factors, and residual platelet function is demonstrated indicating that this product would be expected to be at least equivalent to a single unit of each of the conventional components commonly used in trauma resuscitation.     

Cytokines in WBC-reduced apheresis PCs during storage: a comparison of two WBC-reduction methods

BACKGROUND: Several studies have suggested that cytokine accumulation during storage of platelet concentrates (PCs) may mediate nonhemolytic febrile transfusion reactions and that a reduction in WBC numbers prevents the generation of cytokines. Despite efforts to minimize WBC contamination in apheresis PCs, high numbers of WBCs and increased cytokine levels may still occur, depending on the quality of the apheresis device employed. STUDY DESIGN AND METHODS: This study was undertaken to investigate whether PCs collected with WBC-reduction devices (Spectra LRS, COBE;or MCS+ LDP, Haemonetics) were sufficiently depleted of WBCs to limit cytokine accumulation during storage. The study evaluated 1) the levels of cytokines of WBC and platelet origin in two types of apheresis PCs during storage and 2) the effects of prestorage filtration on cytokine levels in the Spectra LRS PCs. RESULTS: In the Spectra LRS PCs, low levels of IL-6, IL-8, and monotype chemoattractant protein 1 (MCP-1) were detected in Day 1 PCs, and they remained consistent during the shelf life. RANTES, platelet factor 4 (PF4), beta-thromboglobulin (beta-TG), and transforming growth factor (TGF)-beta1 were also detected in these PCs, and their levels increased significantly on storage. Prestorage filtration of Spectra LRS PCs did not further reduce the levels of IL-6, IL-8, MCP-1, PF4, beta-TG, and TGF-beta1 in the filtered component. In the MCS+ LDP PCs, IL-6 was detected on Day 1, and its level increased significantly on storage, whereas the levels in the Spectra PCs remained steady. IL-8 levels were lower in MCS+ LDP PCs than in Spectra LRS PCs of the same age. MCP-1 levels were similar in both products on Day 1 and marginally increased in stored MCS+ LDP PCs. Substantial amounts of RANTES, PF4, beta-TG, and TGF-beta1 occurred in Day 1 MCS+ LDP PCs, and, on storage, these levels rose significantly. CONCLUSION: Despite a significant reduction in levels of WBC-derived cytokines, platelet-derived cytokines were present in different amounts in the two products.     

抗凝全血标本存放条件对血细胞参数稳定性的影响

目的 探讨抗凝全血标本存放条件对血细胞分析仪分析参数稳定性的影响.方法 用Sysmex K-4500血细胞分析仪对EDTA-K2抗凝血标本进行长时间(144 h)不同时段、不同放置温度的血细胞参数稳定性检测,用Excel 软件做统计学分析.结果 室温(18℃~25℃)条件下,白细胞(WBC)可稳定72 h,血小板计数(...     

Estimating efficiency a priori: a comparison of blocked and randomized designs

This technical note deals with a priori estimation of efficiency of functional magnetic resonance imaging (fMRI) designs. The efficiency of an estimator is a measure of how reliable it is and depends on error variance (the variance not modeled by explanatory variables in the design matrix) and the design variance (a function of the explanatory variables and the contrast tested). Changes in the experimental design can induce changes in the variance of estimated responses. This translates into changes in the standard error of the response estimate or equivalently into changes in efficiency. One consequence is that statistics, testing for the same activation in different contexts (i.e., experimental designs), can change substantially even if the activation and error variance are exactly the same. We demonstrate this effect using an event-related fMRI study of single word reading during blocked and randomized trial presentations. Furthermore, we show that the error variance can change with the experimental design. This highlights a problem with a priori comparison of efficiency for two or more experimental designs, which usually assumes identical error variance.     

Rab11B participates in erythrocyte storage lesion of under-collected whole blood

Background and ObjectivesThe storage lesion of the red blood cell affects the life span of RBC and the quality of blood component. The elucidation of this mechanism is helpful to reduce the storage damage of RBC and improve the efficacy and safety of blood transfusion. The aim of this study was to discover the potential molecular mechanism of erythrocyte storage lesion with Under-collected whole blood (UC-WB) model.MethodsThe label-free MS/MS quantitative method was used to identify the differential proteins of erythrocyte membrane proteins and the difference of Rab11B, V-ATPase and plasma GDI2 protein expression were further verified by western blot at the end of blood storage.ResultsA total of 12 Rab proteins and 3 interacting effector proteins were identified among the membrane protein of normal WB and UC-WB, including 5 differential Rab proteins and 2 interacting effector proteins. Compared with normal WB, the expression of membrane Rab11B protein and ATP6V1B1/2 subunit of V-ATPases protein as well as the plasma GDI2 protein of UC-WB increased at the end of storage period.ConclusionRab protein might be related to RBC storage lesions, Rab11B participates in the RBC storage lesion through Rab11B/V-ATPases pathways.     

Controlled storage conditions prolong stability of biochemical components in whole blood.

Blood specimens from primary care centres are normally transported to central laboratories by mail. This necessitates centrifugation and separation, especially since the potassium ion concentration in whole blood changes during storage at ambient temperature. Thus, because of the growing awareness of and concern for pre-analytical contributions to the uncertainty of measurements, we investigated 27 components and their stability under controlled temperature conditions from 17 to 23 degrees C. We found that storage of whole blood can be prolonged by up to 8-12 h for all components examined, including potassium ions, when stored at 20+/-0.2 degrees C. We conclude that this opens the possibility for establishing a pick-up service, by which whole blood specimens stored at 20-21 degrees C can be collected at the doctor's office, making centrifugation, separation and mailing superfluous. In addition, the turn-around time from sample drawing to reporting the analytical result would be shortened. After investments in thermostatted boxes and logistics, the system could reduce costs for transporting blood samples from general practice centres to central laboratories.     

A comparison of medication errors between two storage sites.

This prospective single-blinded study aimed to compare the types of medication errors and medication error rates of two medication delivery systems. The setting was a 30 bed surgical ward that was divided into two identical areas. In one area medications were stored and issued in a ward bay workstation immediately outside the patients' rooms. The alternate area used a medication trolley at the patients' bedside. Three hundred and forty opportunities for errors were observed using five nurse educators. Twenty administration errors (5.8% error rate) and two dispensing errors (0.6% error rate) were detected. A statistically significant difference was found between the two systems, where four errors occurred from the medication trolley (2.6% error rate), and fifteen errors occurred from the ward bay (9.2% error rate). These results suggest that medications were less likely to be omitted and more likely to be given on time when they were issued at the patient's bedside using the medication trolley.     

Potassium load in CPD-preserved whole blood and two types of packed red blood cells

The potassium load of transfused blood must be minimized. We have compared the total plasma potassium content of units of CPD-preserved stored whole blood (SWB), stored packed cells (SPC), and packed cells prepared from stored whole blood (WB-PC). Plasma potassium concentrations, unit weights, and hematocrits of 20 units of SWB, 27 units of SPC, and 20 units of WB-PC of various ages were measured. During the 21-day storage period, total plasma potassium content per unit increased in units of SPC at the same rate as in units of SWB, because plasma potassium concentration increased in SPC at three times the rate of SWB. The values for total plasma potassium per unit at 14 and 21 days in mEq/unit were: SWB, 4.4, 5.8; SPC 3.1, 4.4; and WB-PC 1.9, 2.5. Thus, SPC units may contain substantial amounts of plasma potassium when stored for two to three weeks. However, removal of most of the remaining supernatant plasma from SPC units just prior to administration provides a readily available supply of low potassium blood while allowing maximum conservation of scarce blood resources.