Leucocyte-Depleted Blood: A Comparison of Cell-Washing Techniques

Abstract. Non-haemolytic febrile transfusion reactions are common in multi-transfused patients. It is generally accepted that the majority of these reactions occur when more than 0.5 times 10⁹ leucocytes are transfused. Values equal to or lower than this threshold, which we have called for convenience the critical antigenic load for leucocytes (CALL), can be achieved by decreasing the leucocyte content in one unit of whole stored blood by about 80%. We have compared the efficiency of leucocyte depletion of whole stored blood, using fixed-speed centrifugation (Haemonetics Model 15 and Model 115 cell washer), variable-speed centrifugation (Dideco Progress 90 Cell Separator), and serial centrifugation (IBM 2991 Blood Cell Processor). Fixed-speed centrifugation was least effective in depleting leucocytes; a reduction of 25 and 60% was achieved using the Haemonetics Model 15 and Model 115, respectively. Variable speed and serial centrifugation produced more satisfactory results with leucocyte depletions of 82 and 89% using the Dideco Progress 90 Cell Separator and the IBM 2991 Blood Cell Processor, respectively. Platelet depletion of over 90% was achieved with all the cell washers, except the Haemonetics Model 15. Red cell losses varied from 3 to 30%. It seems unlikely that consistently high (over 90%) leucocyte depletion can be obtained using cell-washing techniques without associated high red cell losses.     

Plateletpheresis Experience with the Haemonetics Blood Processor 30, the IBM Blood Processor 2997, and the Fenwal CS-3000 Blood Processor1,2

Comparisons were made of the apheresis instruments Haemonetics Blood Processor 30, IBM Blood Processor 2997 and Fenwal CS-3000, for collection of platelets from normal donors. With each instrument the mean recovery was at least 4times10¹¹ platelets per procedure, and each instrument afforded a safe and reliable collection. The Haemonetics Blood Processor gave the lowest recovery of platelets per minute per procedure. The IBM Blood Processor 2997 required the longest time for set-up and priming and processed 1.5 liters more donor blood per collection than the other instruments. The Fenwal CS-3000, which is a computer-controlled instrument, was the least time consuming. The donor suffered a significantly greater drop in platelet count after collection with the IBM Blood Processor 2997 (31%) than after collection with the other instruments (19%), and we were unable to account for this observation.     

A New Approach to Washing Red Blood Cells Frozen with a High Concentration of Glycerol in a Special Freezing Container1

We report here on a new approach to washing red blood cells frozen with a high concentration of glycerol in a special freezing container. The wash solution consists of a 150-ml volume of 12% sodium chloride and 2 liters of 0.9% sodium chloride-0.2% glucose-25 mEq/l disodium phosphate. Both the Haemonetics Blood Processor 115 and the IBM Blood Processor 2991 have been used with this protocol, with similar results. The in vitro recovery of red blood cells frozen with 8.6M glycerol was 89%, and that of red blood cells frozen with 6.2M glycerol was 93%. The 24-hour posttransfusion survival values averaged 88% for eight units of outdated-rejuvenated previously frozen red blood cells washed by this protocol and stored at 4°C for 3 days before autotransfusion.     

A Technique for the Concentration of Nucleated Bone Marrow Cells for in vitro Manipulation or Cryopreservation Using the IBM 2991 Blood Cell Processor

Abstract. 27 bone marrow aspirates were processed on the IBM 2991 Blood Cell Processor to achieve a reduction in volume (>80%) and in red blood cell contamination (>75%), without loss (<20%) of nucleated cells. The procedure was used to concentrate nucleated bone marrow cells either prior to cryopreservation for subsequent autologous transplantation, or prior to incubation with a murine monoclonal antibody (OKT3) for allogeneic transplantation. We conclude that the procedures used for the concentration, cryopreservation, thawing and infusion do not adversely affect the viability of cells as assessed by in vitro culture (CFU-GM). Of the marrows processed, 12 have been reinfused and resulted in successful engraftment.     

Standardization of the Processing of Human Bone Marrow for Allogeneic Transplantation

Allogeneic bone marrow transplantation has been undertaken within this centre on 62 patients with acute or chronic leukaemia. Employing the standard separation protocol described, all bone marrows were processed on the 2991 Blood Cell Processor to isolate the 'buffy coat' cells and subsequently the mononuclear cell component using a density separation medium of Ficoll-metrizoate. Following the mononuclear cell separation, the cells were identified for their T lymphocyte component using a combination of two murine monoclonal antibodies, MBG6 reacting with a pan T cell antigen (CD6) and RFT8 detecting the 'cytotoxic/supressor' cell antigen (CD8). The numerical results for nucleated cells, red blood cells, T lymphocytes and colony forming units-granulocyte macrophage are presented.     

A simple technique for preparation of young red cells for transfusion from ordinary blood units

We have developed a rapid and simple method for the isolation of relatively young cells from ordinary fresh or previously frozen blood units, using the IBM-2991 Blood Cell Processor. Based on in vitro measurements of red cell pyruvate kinase activity, the resolution of the method is excellent, comparable to that of the stractan discontinuous gradient system. In splenectomized patients with thalassemia major, the administration of the "youngest" 50% of blood units yielded 51Cr survival times of 40 and 42 days for fresh packed red cells and 41 and 43 days for previously frozen red cells.     

The Utilisation of a New Strength Citrate Anticoagulant during Centrifugal Plateletpheresis

Employing the Haemonetics Blood Processor (IFC), a relatively pure platelet concentrate can be prepared by collecting only the first portion of the PRP leaving the centrifuge bowl (Fraction I). A subsequent fraction containing RBC and WBC contaminants (Fraction II) can be purified by means of a second centrifugation, using a conventional blood bank centrifuge (Fraction II), if transfusion of these contaminants would be detrimental to the recipient. Utilising the new 1.4% Na3-citrate anticoagulant, platelet metabolic parameters (ATP, ADP, AMP, lactate and pyruvate) as well as O2-uptake, were determined in Fraction I and II prepared from 10 normal healthy subjects. In contrast to previous studies reporting marked dysfunction in platelets contained in Fraction II when standard ACD-A was used during IFC, we observed no significant difference (Student's t test) in the present study between Fractions I and II, in regard to platelet metabolism, when using the new anticoagulant. It is further concluded that the second centrifugal manipulation does not exert a detrimental effect on platelet metabolism.     

Blood pressure reactivity to psychological stress and coronary calcification in the Coronary Artery Risk Development in Young Adults Study

A longstanding hypothesis is that individuals who exhibit large increases in blood pressure during psychological stress are at risk for atherosclerosis. We tested whether blood pressure changes during psychological stress predict subsequent coronary calcification (CaC) in young healthy adults. We evaluated 2816 healthy black and white women, 20 to 35 years of age, from the Coronary Artery Risk Development in Young Adults Study, who were not using medication for hypertension or diabetes in 1987-1988. Participants completed video game and star tracing tasks while their blood pressure was recorded. Thirteen years later (2000-2001), they completed computed tomography measures of CaC. Overall 9.3% (261 of 2816) had CaC present at follow-up. Each 10 mm Hg change in systolic blood pressure during the video game was associated with a 24% increased odds of having CaC at follow-up (unadjusted odds ratio, 1.24; 95% CI, 1.06 to 1.46; P=0.008). This association persisted after adjustment for age, race, sex, education, smoking, alcohol, family history of myocardial infarction, smoking, daily alcohol consumption, body mass index, and resting or baseline blood pressure (odds ratio, 1.31; 95% CI, 1.08 to 1.58; P=0.006). Blood pressure changes during the star tracing task were not associated with subsequent CaC. Blood pressure changes during a video game predicted the presence of CaC 13 years later. To our knowledge, this is the first study that reports blood pressure reactivity to a stressor being related to calcification in the coronary arteries. Blood pressure reactivity may provide useful prognostic information about future risk beyond standard risk factors.     

Influence of Antioxidants on the Quality of Stored Blood

Background and objectives: Blood is exposed to oxidation stress and therefore has a high antioxidant capacity (AOC). With the many factors increasing the demands on the AOC, there may be damage to erythrocytes by free radicals. This study was to investigate evidence of erythrocyte damage in stored donor blood and to affect this by premedication of blood donors. Materials and methods: Blood samples of 15 healthy donors were collected in CPDA-1 solution and analyzed immediately, and then again after 10 days of incubation at 4 °C and 1 day of incubation at 37 °C. Prior to incubation, the following parameters were evaluated: Na⁺, K⁺, malondialdehyde (MDA), hemoglobin (Hb), AOC in the supernatant, superoxide dismutase (SOD) in erythrocytes, and glutathione peroxidase (GSHPx) in whole blood. Blood donors of group 1 were not given any drugs or vitamins before blood sampling. The same blood donors were then supplemented with the following daily doses of antioxidants for 10 days before the next blood sampling: 36 mg of β-carotene, 300 mg of vitamin E, 200 mg of vitamin C, and 40 mg of selenium. Results: The blood from donors of group 2 had a significantly smaller increase in MDA, K⁺, and Hb, and a smaller decrease in Na⁺ and AOC in the supernatant compared with that of group 1, while the activity of SOD and GSHPx did not change during blood storage. Conclusions: These results suggest that antioxidants given to blood donors can improve red cell storage parameters by reducing cell damage caused by free radicals.     

Contributions to the Optimal Use of Human Blood

The influence of different variables on the yield of factor VIII in cryoprecipitate as prepared in the Central Laboratory of the Netherlands Red Cross Blood Transfusion Service, was studied. The following conclusions may be drawn: (1) In case blood should be stored, the use of the anticoagulant solution acid citrate dextrose (ACD) is preferable to citrate phosphate dextrose (CPD) or trisodium citrate (TSC). (2) The temperature of stored whole blood should not decrease below 8 degrees C because of a spontaneous precipitation of factor VIII from blood (and plasma) below this temperature. (3) Cryoprecipitate derived from rapidly frozen plasma (1 min) contains a decreased amount of proteins in comparison with cryoprecipitate prepared from slowly frozen plasma (45 min to 4 h). On the other hand, equal amounts of factor VIII activity were obtained in the precipitate after freezing of plasma at varying rates. (4) Rapid thawing of plasma results in both higher yields of factor VIII procoagulant activity and a higher specific activity of this factor in the resulting cryoprecipitate. (5) The sedimentation of cryoprecipitate is completed after 5 min centrifugation at 1,500 g. (6) At temperatures higher than 8 degrees C, cryoprecipitated factor VIII starts dissolving into the supernatant plasma or in buffer. (7) Factor VIII in lyophilized cryoprecipitate is stable at room temperature. At elevated temperatures it rapidly looses its activity. (8) Evidence was obtained that the improvements which are introduced in the preparation of factor VIII do not lead to a product which is less stable in vitro as well as in vivo.     

Comparison of blood rheological changes in the microcirculation during experimental hemorrhagic and traumatic shock

OBJECTIVE: The aim of the present study was the analysis of the specific hemorheological disorders during the middle stages of experimental traumatic and hemorrhagic shock. METHODS: The following hemorheological properties were investigated: Red Blood Cell (RBC) aggregability, their deformability, and systemic hematocrit. The RBC aggregability was assessed by using the "Georgian technique". The RBC deformability was determined with the nuclepore membrane filter method of Reid. The hematocrit was measured by blood centrifugation. The results obtained were treated statistically by using the "Two sample T-test". RESULTS: We found that in the animals with traumatic shock the erythrocyte aggregability index increased by mean 181%, while in the hemorrhagic group this index on contrary decreased by mean 68% as compared to control group; The RBC deformability underwent a significant decrease during both traumatic and hemorrhagic shock: by a mean of 52% during the first and 62% during the second one. The systemic hematocrit decreased by a mean of 45% during the traumatic shock and by a mean of 50% during the hemorrhagic shock. CONCLUSION: The data obtained provide evidence that the hemorheological disorders are among the most significant microcirculatory disturbances in the pathogenesis of both the traumatic and the hemorrhagic shock.     

Rate of Seroconversion in Repeat Blood Donors at The National Blood Centre,Kuala Lumpur

The World Health Organization (WHO) recommend that all donated blood are to be screened for at least three viral infections [human immunodeficiency virus (HIV), hepatitis C virus (HCV), hepatitis B virus (HBV)]. The National Blood Centre, Kuala Lumpur (NBCKL) aims to reduce transfusion transmitted infections (TTI) as it still remains as one of the major risk for blood transfusion. A cross sectional study was conducted at the National Blood Centre, Kuala Lumpur from 1st January 2009 to 31st June 2010. Data from 581,020 donors were analyzed from year 2004 to 2008. All data were retrieved from NBCKL Blood Bank Information System (BBIS). A total of 201 repeat donors were included in the study based on the inclusion criteria but only 132 repeat donors agreed to participate. Information on sociodemographic, risk factors, knowledge of donors and high risk behavior were extracted from standardize questionnaire. Data were analyzed using SPSS version 14.0. The aim of this study was to determine the predictors of the seropositive infectivity among repeat blood donors at the NBCKL. The results showed Syphilis accounts for the highest and increasing seroconversion rate among other infections from 20.83 % in year 2004 to 44.6 % in year in year 2008. HIV and HCV infection also showed increasing seroconversion rate in 5 years’ time from 6.41 % in year 2004 to 17.54 % in year 2008 and 4.8 % in year 2004 to 5.94 % in year 2008 respectively. However, HBV infection alone showed a decreasing seroconversion rate from 20.83 % in year 2004 to 10.4 % in year 2008. Level of donors’ awareness regarding high risk factors (activities or behaviour) can lead to higher risk of TTI with significant p value in this predictors model(p < 0.05). Repeat blood donors with high risk activities are more likely to have seropositive results for HBV, HIV and Syphilis. This study found that the frequency of HCV seropositivity is higher among lapsed donor. Socio demographic factors such as male and working in the private sector predominates in all TTI markers. Majority of the respondents were aware about relation of high risk activities and risk of TTI.     

Granulocyte Yields Using the Haemonetics 30

Abstract. The factors of donor selection, corticosteroid drug choice, dose, timing and route of administration were studied to optimize granulocyte yield employing the Haemonetics Model 30 Blood Processor. Our data suggest that donors giving one excellent yield do not necessarily do so again. Donors with high initial granulocyte counts had only marginal increases in counts after taking corticosteroids. Selecting donors on the basis of high previous yield or granulocyte count is not a feasible strategy for improving granulocyte yields. Highest yields of granulocytes (16times1o⁹ total; 4.1times10⁹/liter processed) were obtained with any of four split-dose corticosteroid regimen. This increment is almost twice as large as the previously reported improvement using continuous-flow centrifugation. The choice of steroid, the route of administration and the timing of the split doses are not critical.     

The Donation Cycle: A Framework for the Measurement and Analysis of Blood Donor Return Behaviour

The donation cycle represents a new framework for the measurement of blood donor return behaviour. Because it is based on the interval between successive donation attempts, it is more efficient than previously reported methods and the resulting data can be analysed using the statistical techniques for interval data. To illustrate the merits of this quantitative approach to the study of blood donor behaviour, the donation cycle framework is used to analyse the interval between the first and second donation attempts in a random sample (n = 5,183) of type 0, whole blood donors from the Gulf Coast Regional Blood Center. Simple statistical tools such as the log-rank test are employed to describe and to evaluate relative differences in the return behaviour of Rh-negative and Rh-positive donors. The analysis indicates that Rh-negative donors are significantly (p < 0.001) more likely than Rh-positive donors to attempt to donate on a second occasion. Kaplan-Meier estimates of the survival function of these first-time donors reveal marked elevations in the rate of return exactly 52 weeks after the initial donation. The donation cycle paradigm provides transfusion researchers with quantitative tools which are essential for designing statistically efficient, prospective intervention studies. By using the knowledge which such studies could provide, blood banks might be better able to manage donor return behaviour and thereby the safety of the blood supply.     

Contributions to the Optimal Use of Human Blood

Abstract . Analysis of the production of plasma protein fraction (PPF) showed a considerable loss of albumin (18 to 25% of the amount present in whole blood) in the cell concentrate after centrifugation of blood and in fraction IV (11% of the amount present in plasma) during ethanol fractionation with a modified Cohn VI method.
The recovery of albumin could be increased by washing 10 volumes of cell concentrate, derived from blood centrifuged in a 'bottle centrifuge' with one volume of 6% NaCl solution. After centrifugation, 120 ml supernatant per unit of blood is obtained with a protein content of 3%. This diluted plasma is mixed with normal plasma and fractionated according to the modified Cohn VI method. The effect of the described modifications is that the quantity of PPF obtained from blood centrifuged in a 'bottle centrifuge' is at least 30% more, not only by the recovery of the plasma proteins formerly remaining in the red cell suspension, but also by a decreased loss of albumin in fraction IV. The final product meets the minimum requirements for PPF.
When the blood is centrifuged in a continuous centrifuge, the loss of albumin could be decreased from 18 to 5% by using 2.6% trisodiumcitrate and 0.72% NaCl as an anticoagulant solution instead of ACD and by a lower flow rate.     

Blood volume measurement as a tool in diagnosing syncope

BACKGROUND: The cause of syncope remains unknown in 24% to 37% of cases even after standard diagnostic tests. Measuring blood volume may elucidate the mechanisms of syncope in the individual patient and prove helpful in determining optimal therapy. This report includes the largest set of blood volume measurements performed in syncope patients to date. METHODS: We performed radioisotopic blood volume measurement (Daxor BVA-100) on 539 patients who presented to our center with syncope/presyncope of unclear etiology. There were 202 men and 337 women, ages 16 to 88 years; many were receiving treatment at the time they were referred to our center. We also measured blood pressure, heart rate, and cardiac index before and during tilt, with complete data available for 411 patients. RESULTS: Blood volume derangements ranged from -32% to +116% deviation from normal. Hematocrit could not be used to predict volume status. Volume depletion was found in 241 (44.7%) patients and volume expansion was found in 63 (11.7%). Blood pressure, heart rate, and cardiac index before and during tilt did not correlate with any component of blood volume and could not be used to predict volume status. CONCLUSIONS: Syncope patients are heterogeneous with respect to blood volume, and blood volume derangements are common and are not identified through tilt table testing. Empirically prescribed pharmacological treatment for syncope is frequently inappropriate. Blood volume measurement should be included in syncope diagnosis.     

Clarifying the Relationship Between ABO/Rhesus Blood Group Antigens and Upper Gastrointestinal Bleeding

Background and Aim The relationship between blood group antigens and peptic ulcer disease has been widely evaluated in the past. Data concerning the same association with upper gastrointestinal bleeding are very limited. We aimed to evaluate this association and we thought it was worthwhile to try to determine whether these components take some part in this complication. Methods The study population consisted of 1,098 adults (364 patients and 734 volunteer blood donors as controls). Demographic features, comorbid illnesses, and use of aspirin/nonsteroidal anti-inflammatory drugs (NSAIDs) were recorded. Blood groups were examined by gel centrifugation method. We included only patients with bleeding from peptic ulcer disease and erosive gastropathy. Ulcers were classified by using Forrest’s classification system in terms of rebleeding risk. Helicobacter pylori was examined by histology. Results The gender distribution was similar in both groups. The ABO blood group phenotype distribution in patients and controls (respectively) was as follows: 46.2% versus 34.9% for group O, 32.4% versus 39.5% for group A, 15.7% versus 18.4% for group B, and 5.8% versus 7.2% for group AB. Blood group O was found to have higher frequency in the patient group than in the control group (P = 0.004). Rh positivity was also higher in patients than in controls (P = 0.007). H. pylori positivity was similar between blood groups among patients. The rebleeding and mortality rates between blood groups were also similar. Conclusion ABO blood group O had an important role in patients with upper gastrointestinal bleeding. The impact of blood group on rebleeding and mortality may be a focus for further studies.     

高血压病人动脉顺应性与家庭偶测血压及动态血压关系

目的　探讨原发性高血压病人家庭测量血压、动态血压与动脉顺应性的关系。方法　共入选 164例受试者 ,其中原发性高血压组 74例 [男 42例 ,女 3 2例 ,平均年龄 ( 4 7 0 5± 10 0 8)岁 ] ;对照组 90例 [男 41例 ,女 49例 ,平均年龄 ( 3 3 81±12 3 5 )岁 ]。对入选者间隔 1～ 3周进行二次家访 ,每次家访由经过专门训练的护士采用汞柱血压计连续测量受试者坐位、左上臂血压 5次 ,每人的家庭测量血压是二次家访、共 10个血压读数的平均值。同时采用SpaceLabs 90 2 0 7动态血压监测仪记录 2 4小时动态血压 ,有效数据应达到 80 %以上。动脉顺应性检查采用动脉脉搏波分析 (PWA)仪。反映大动脉硬化的参数为反射波增强指数 (AIx)。结果　高血压组家庭测量血压及 2 4小时平均收缩压、舒张压、白昼及夜间血压显著高于对照组。与对照组相比 ,高血压组AIx显著升高 ( 2 4 4%± 12 8%vs 12 7%± 18 3 % ,P =0 0 0 0 ) ,经年龄、性别调整后 ,二组间仍有显著性差异 ,提示高血压患者动脉顺应性下降、动脉硬化。将AIx作为因变量 ,年龄、身高、家庭测量血压及动态血压中 2 4小时平均血压、白昼血压、夜间血压、血糖、血酯等作为自变量进行多元回归分析 ,AIx与年龄、家庭血压中的收缩压呈显著的正相关 ,与身高呈负相关     

Blood D-3-hydroxybutyrate and the regulation of plasma concentrations of free fatty acids in the fasted rat.

Acetoacetate or DL-3-hydroxybutyrate were infused into fasted rats at rates comparable to the endogenous rate of ketone body release. Blood samples taken before and during infusion were analyzed for concentrations of ketone bodies, free fatty acids, and immunoreactive insulin. During infusion of acetoacetate or DL-3-hydroxybutyrate, blood concentrations of total ketone bodies doubled. While the HBA/AcAc ratios remained constant in rats infused with DL-3-hydroxybutyrate, the ratios decreased significantly during infusion of acetoacetate, reflecting increases in acetoacetate concentrations without proportionate increases in concentrations of D-3-hydroxybutyrate. Blood concentrations of free fatty acids decreased more rapidly in the rats infused with DL-3-hydroxybutyrate than in the rats infused with acetoacetate. The decrease in free fatty acid concentrations was related to increases in blood D-3-hydroxybutyrate in both the DL-3-hydroxybutyrate-infused rats and in the acetoacetate-infused rats. Plasma immunoreactive insulin concentrations were significantly higher in the DL-3-hydroxybutyrate-infused rats than in the acetoacetate-infused rats. Acetoacetate and DL-3-hydroxybutyrate were infused into depancreatized, functionally hepatectomized fasted rats at rates comparable to those used in the intact fasted rats. Only small decreases in free fatty acid concentrations were observed during infusion. From these experiments it was concluded that increases in blood concentrations of D-3-hydroxybutyrate were more effective than acetoacetate in causing decreases in free fatty acid concentrations in the fasting rat. The pancreas is required for this effect.     

Blood flow through the brachial artery at different temperatures in patients with Raynaud's phenomenon

Blood flow through the brachial and radial artery at different temperatures and during reactive hyperaemia was measured in 23 patients with Raynaud's phenomenon (9 patients with Raynaud's disease, 7 - Raynaud's syndrome with thoracic outlet syndrome (TOS), and 8 - Raynaud's syndrome and scleroderma), and 11 control subjects. Blood flow was determined by an ALVAR pulse-wave Doppler system enabling measurement of vessel diameter and blood flow rate. The measurements suggest that the blood flow through brachial artery reflects the changes of blood flow through the hand immersed in water of different temperatures. Maximum vasoconstriction was the same in all groups of Raynaud's phenomenon. The lowest blood flow at higher temperatures and during reactive hyperaemia was found in patients with scleroderma, whereas in the group of TOS and partly in patients with Raynaud's disease one can suspect only functional changes.