bluuebunny/crossref_metadata_2025 huggingface dataset:
Current Status of Hepatitis G Virus (GBV‐C) in Transfusion: Is It Relevant?
Authors: Karayiannis P. , Thomas H.C.
Published: December 2004

A new flavivirus, provisionally designated hepatitis G virus or GBV‐C has recently been described. The virus is parenterally transmitted by exposure to blood through transfusion, intravenous drug use and haemodialysis. Heat‐ or chemically‐treated blood products are associated with reduced risk of infection. The virus may also be transmitted from mother to infant and by the sexual route. Although hepatitis G virus has been detected in patients with acute and chronic hepatitis, a causative role of the virus in such cases has not been established. The majority of long term carriers of the virus appear to have no liver, biochemical or histological abnormalities.
Audit of Practice in Platelet Refractoriness
Authors: Phekoo K.J. , Hambley H. , Schey S.A. , Win N. , Carr R. , Murphy M.F.
Published: December 2004

Objectives: A significant number of patients become refractory to platelet transfusion and prompt investigation of the cause will encourage appropriate selection of platelet products. Methods: We surveyed haematologists to assess perceived practice concerning platelet refractoriness because of the high cost and limited availability of HLA‐compatible platelets. Some 56 of 58 consultant haematologists participated. Results: Clinicians differed on their definition of platelet refractoriness, and non‐immune factors were not considered as important as immune causes of platelet refractoriness. A working group, including an invited moderator, was established to produce guidelines on recommended practice for the management of platelet refractoriness. Re‐audit after implementation of the guidelines showed that more patients receiving HLA‐compatible platelets had been tested for HLA antibodies. There was a mean 50.9% reduction in the use of HLA‐compatible platelets. Conclusions: Increased testing for leucocyte and platelet antibodies resulted in reduced demand for and more selective use of HLA‐compatible platelets, with no apparent increase in haemorrhagic complications.
Examining Technical Aspects of the Monoclonal Antibody Immobilisation of Granulocyte Antigen Assay
Authors: Minchinton Robyn M. , Noonan Kevin , Johnson Tamzine J.
Published: December 2004

Background and Objectives: This study was an attempt to improve detection and characterization of alloantibodies to neutrophil antigens, using the monoclonal antibody immobilisation of granulocyte antigens (MAIGA) assay. Materials and Methods: We explored the effect of different detergent concentrations, detergent class, stabilising additives, combinations of detergents, and use of a broad range of monoclonal antibodies on the results of the MAIGA assay. Results: Non‐ionic detergents, Triton X and Nonidet, were confirmed as the most suitable for solubilisation of the recognised, neutrophil‐associated alloantigens. Anionic and zwitterionic detergents did not assist the demonstration and characterisation of new neutrophil alloantigens in the MAIGA assay. Some reproducible positive MAIGA assay results were obtained for antisera 01, 08, anti‐RED, anti‐NB2, and anti‐NB1, using particular monoclonal antibodies. Conclusions: Although we are reluctant to suggest association of the targeted antigens with the particular glycoproteins identified by the capture monoclonal antibodies, NB1 glycoprotein may be closely associated, before or after antibody binding, with CD46, CD11b, CD50, and/or CD 66b.
Factor VII and Activated‐ Factor‐VII Content of Prothrombin Complex Concentrates1
Authors: Hellstern Peter , Beeck Hannelore , Fellhauer Annette , Fischer Antje , Faller‐Stöckl for the PCC Study Group Beate
Published: December 2004

Background and objectives: The aim of this study was to determine the potencies of factor VII (FVII) and of activated FVII (FVIIa) in prothrombin complex concentrates (PCC). Materials and methods: We examined 56 lots of PCC from 5 manufacturers. Three brands were licensed preparations, and 1 product series had been involved in thromboembolic complications. FVII and FVIIa were measured using a two‐stage amidolytic assay and a specific clotting assay, respectively. We also quantified FVII clotting activity by a one‐stage assay reflecting a mixture of FVII zymogen and FVIIa. Results: All PCC contained substantial amounts of FVII, and FVIIa could be detected in all lots. There were marked differences between manufacturers and some significant variabilities between batches. The two lots involved in thromboembolic events contained considerably more FVIIa than the PCC still licensed. The lowest FVIIa potencies were observed in an experimental product series, indicating that PCC can be produced without activation of FVII during the manufacturing process. Conclusion: FVIIa is present in all PCC containing FVII. High FVIIa potencies may contribute to the thrombogenic potential of these preparations, and determination of FVIIa potencies should be included in the in vitro characterization of PCC.
Moving Blood
Authors: Pelis Kim
Published: December 2004

Our internationally acclaimed journalist Sanguinia has returned safely from her historic assignment. Travelling from Homeric Greece to British Romanticism, she was witness to blood drinking, letting, bathing, and transfusion. In this report, she explores connections between the symbolic and the sadistic; the mythic and the medical – all in an effort to appreciate the layered meanings our culture has given to the movement of blood between our bodies.
Clinical and Laboratory Experience with Erythrocyte and Platelet Preparations from a 0.5CPD Erythro‐Sol Opti System
Authors: Högman C. F. , Eriksson L. , Wallvik J. , Payrat J. M.
Published: December 2004

Background: Red blood cells stored as concentrates or suspensions in additive solutions change rapidly their oxygen affinity mainly due to the loss of 2,3‐diphosphoglycerate (2,3‐DPG). When collected in CPD with half of the normal concentration of citrate and citric acid (0.5CPD) and stored in a new additive solution (Erythro‐Sol), 2,3‐DPG is better maintained. No studies of the oxygen affinity of red cells stored under these conditions have been published. In Erythro‐Sol, red cells have a satisfactory in vivo recovery for 49 days but the conditions after 28 days, within which time most red cell units are transfused, have not been investigated. Of importance is also to be able to make platelet concentrates (PCs) from 0.5CPD blood. Little data are available concerning the clinical usefulness of platelets prepared from 0.5CPD buffy coats (BCs). Methods: Blood was collected in 0.5CPD, held at 20°C for 3–4 h, then separated with the bottom‐and‐top technique into red cells, plasma and BC. In a storage experiment with 6 U the 2,3‐DPG and P50 values were determined weekly and a number of in vitro parameters were tested on day 28. In 6 donors the in vivo recovery and survival of red cells were determined using a single‐chromium technique. Transfusions of 212 0.5CPD‐Erythro‐Sol red cell units were given to hematological patients under supervision. PCs derived from pools of 0.5CPD BCs suspended in PAS2 (T‐Sol) were transfused to 20 thrombocytopenic patients and compared with CPD‐BC‐PCs suspended in PAS1. Corrected count increments (CCI) were determined. Results: The erythrocyte 2,3‐DPG and P50 values were normal or slightly subnormal initially but increased to supernormal levels during the 1 week, and remained at these levels for a further 1–3 weeks; the 2,3‐DPG was two thirds of normal after 28 days, the P50 was 3.72±0.28 kPa after 14 days and 2.84±0.41 after 28 days (mean ± SD). The P50 values corresponded closely (r2= 0.903) to 2,3‐DPG. The in vivo recovery of 4‐week‐stored red cells was 89.6±5.5% and the T50 was 32.2±2.0 days. No adverse effects were observed in the transfusions. The CCI values did not differ between test and control groups; in both, 3‐ to 5‐day‐stored PCs gave lower CCI than fresh (0–2 days) PCs. Patients with acute myeloid leukemia AML (n = 11) had significantly lower CCI values than patients with myelodysplastic syndrome, myeloma and lymphoma (n = 9; CCI1 h: p = 0.001; CCI24 h: p = 0.006). Conclusions: Red cells stored in Erythro‐Sol sustain a normal or slightly lowered oxygen affinity for 2–4 weeks, their viability is excellent, and they are well tolerated in clinical transfusions. Platelets prepared from 0.5CPD‐BCs cause CCls of the same magnitude as CPD‐BCs.
Evaluation of Two Different Protocols for Peripheral Blood Stem Cell Collection with the Fresenius AS 104 Blood Cell Separator
Authors: Menichella Giacomo , Lai Marco , Pierelli Luca , Vittori Mariangela , Serafini Riccardo , Ciarli Marina , Foddai Maria Laura , Salerno Giovanna , Sica Simona , Scambia Giovanni , Leone Giuseppe , Bizzi Bruno
Published: December 2004

Objectives: Reconstitution of hematopoiesis by means of peripheral blood stem cells is a valid alternative to autologous bone marrow transplantation. The aim of this investigation was to increase the efficiency of collection of circulating blood progenitor cells and to obtain a purer product for transplant. Methods: We carried out leukapheresis procedures with the Fresenius AS 104 blood cell separator, using two different protocols, the previously used PBSC‐LYM and a new mononuclear cell collection program. Results: Both programs were highly effective in collecting mononuclear cells (MNC) and CD34+ cells. Some differences were found, especially regarding MNC yield and efficiencies. There are remarkable differences in the efficiency of collection of CD34+ cells (62.38% with the new program as opposed to 31.69% with the older one). Linear regression analysis showed a negative correlation between blood volume processed and MNC efficiency only for the PBSC‐LYM program. Differences were also observed in the degree of inverse correlation existing in both programs between patients' white blood cell precount and MNC collection efficiency. The inverse correlation was stronger for the PBSC‐LYM program. Seven patients with solid tumors and hematologic malignancies received high dose chemotherapy and were subsequently transplanted with peripheral blood stem cells collected using the new protocol. All patients obtained a complete and stable engraftment with the reinfusion product collected with one or two leukapheresis procedures. Conclusions: High efficiencies and yields were observed in the new protocol for MNC and CD34+ cells. These were able to effect rapid and complete bone marrow recovery after myeloablative chemotherapy.
The Serological Profile and Molecular Basis of a New Partial D Phenotype, DHR
Authors: Jones J.W. , Finning K. , Mattock R. , Williams M. , Voak D. , Scott M. L. , Avent N. D.
Published: December 2004

Background and objectives: The Rh D antigen comprises a mosaic of at least 30 epitopes expressed on a 30‐kD non‐glycosylated Rh D polypeptide. The equivalent Rh CeEe polypeptide expressing the Rh C/c and E/e antigens differs in only 36 of the 417 amino acid residues. Partial D individuals have been described who fail to express a number of D epitopes. Materials and methods: Serologic methods were applied with monoclonal anti‐D to map epitopes on the red cells of a proposita aberrant D typing. Polymerase chain reaction (PCR) and DNA sequencing were also done. Results: DNA sequence analysis derived by RT‐PCR using total RNA isolated from peripheral blood of this person suggests two mechanisms for the genetic basis of this variants: one where gene conversion events result in the replacement of RHD gene exons with the equivalent RHCE exons; the second where point mutation in the RHD gene generates an amino acid substitution in the Rh D protein. Conclusions: We report here a new partial D, DHR, where a single point mutation (G to A at nucleotide 686) in exon 5 of the RHD gene results in a conservative amino acid substitution (Arg229Lys), in the predicted Rh D protein. This residue is localised on the fourth predicted exofacial loop of the Rh D polypeptide as determined by hydropathy analysis. This substitution results in the lack of epD 1, 2, 12 and 20 (30 epitope model) and indicates the involvement of loop 4, and in particular the requirement of Arg229, in the expression of these epitopes.
IgA Content of Washed Red Blood Cell Concentrates
Authors: Tóth C. Baróti , Kramer J. , Pinter J. , Thék M. , Szabó J.E.
Published: December 2004

Saline‐washed red blood cell (RBC) concentrates are largely depleted of leukocytes, microaggregates, and nearly of all plasma originally contained in the blood unit. Our aim was to determine the concentration of the eliminated IgA and total protein in the buffy coat and plasma removed RBC concentrate before and after a sequential dilution‐centrifugation washing procedure. 75.3% protein, and 99% IgA were eliminated by the washing process used. The recovery of red blood cells was 87.2%. Our results have led to the conclusion that two sequential washings seem to guarantee an IgA level of less than 0.2 mg/unit to meet the recommendations of the Council of Europe. Additional advantages of washing include low levels of extracellular hemoglobin, metabolic waste products and debris in the supernatant.
Complement Activation during Blood Sampling Procedures Alters the Expression of CD11b/CD18 on Human Neutrophils
Authors: Wehlin L. , Gustavsson K. , Halldén G. , Emilson A. , Svensson A. , Hild M. , Lundahl J.
Published: December 2004

Background and objectives: Differences in blood sampling and separation techniques can affect the quantitative levels of activation markers on different leukocyte subsets. We examined the effect of two sampling procedures of EDTA blood on the quantitative levels of two markers, the CD11b/CD18 antigen and the EG2 epitope on intracellular eosinophilic cationic protein (ECP), in neutrophils and eosinophils, respectively. Materials and methods: Sample I was collected directly after completion of blood donation by an open technique and constant flow from the transfer tube directly into EDTA tubes. After sampling, the transfer tube was manually closed with a clamp. Sample II was collected 45 s later by the same technique by opening the clamp. Results: We found a significantly (p < 0.01) higher expression of CD11b/CD18 on neutrophils collected by sampling procedure II than on those collected by sampling procedure I. In contrast, we did not find any difference in the intracellular ECP expression between sampling procedures I and II. To further explore the mechanisms for the observed upregulation of CD11b/CD18, fragments of a transfer tube were incubated with normal human serum (NHS) and heat‐inactivated NHS (NHS56), respectively, for 60 min at +37oC. Leukocytes from healthy blood donors were then incubated for 15 min at +37oC with these serum preparations. The CD11b/CD18 expression was significantly higher (p < 0.01) on neutrophils incubated with transfer‐tube‐activated NHS compared with NHS alone. However, when leukocytes were incubated with transfer tube activated NHS56, no difference was observed compared with incubation with NHS alone. In addition, by using confocal laser scanning microscopy, we could identify complement (C3c) deposits on the inner surface of the transfer tube fragments incubated in NHS, but not in NHS56. Conclusions: The quantitative level of the activation marker CD11b/CD18 on neutrophils, but not the EG2 epitope on intracellular ECP in eosinophils is significantly increased by a slight modification of the blood sampling procedure. It is suggested that the observed upregulation of CD11b/CD18 is caused by complement activation within the transfer tube. The results emphasize the importance of in‐house data on the effect of variations in sampling procedures, particularly when data from healthy blood donors are included in clinical studies.
Flow Cytometry Immunophenotyping and Polymerase Chain Reaction‐Site‐Specific Primers Genotyping for HPA‐1 Alloantigens in an Italian Blood Donor Population
Authors: Tazzari Pier Luigi , Cirillo Dora , Bontadini Andrea , Ricci Francesca , Masi Renato , Conte Roberto
Published: December 2004

Background and objectives: There is increasing interest in the development of rapid and reliable techniques for human platelet alloantigen (HPA) typing. This study investigates the reliability of flow cytometry for large‐scale immunophenotyping of platelet alloantigens. Materials and methods: We used flow cytometry and polymerase chain reaction‐site‐specific primer (PCR‐SSP) for the characterization of the human platelet antigen 1 (HPA‐1) mosaic in blood donors. Results: By using specific alloantisera and immunofluorescence labelling 9 (2.6%) out of 351 samples were HPA‐la‐negative. To confirm this antigenic phenotype, all of the latter samples were submitted to PCR‐SSP analysis, showing an HPA1‐b/b genomic pattern. In HPA‐la‐positive donors, flow cytometry was unable to distinguish HPA‐1a/b heterozygous from HPA‐1a/a homozygous subjects who were clearly identified by genotyping. Conclusions: Flow cytometry is a valuable tool for large‐scale screening to identify HPA‐la‐negative persons, whereas genotyping is the assay of choice for zygosity testing, antenatal diagnosis, and for thrombocytopenic alloimmunized patients.
Heterogeneity of the O Alleles at the Blood Group ABO Locus in Amerindians
Authors: Olsson Martin L. , Santos Sidney E. B. , Guerreiro Joao F. , Zago Marco A. , Chester M. Alan
Published: December 2004

Background and objectives: Amerindians are blood group O, but the distribution of the various O alleles is unknown. Their ABO genotypes were compared with samples from other Brazilian ethnic groups. Materials and methods: Genomic DNA was examined by PCR‐RFLP analysis, PCR‐SSP and direct sequencing. Results: An unusual allele distribution was found, with 91% of the O alleles being OIvariant. Almost half of these alleles had an additional novel mutation (G542A), which was also detected in a few other Brazilian and European samples. The O alleles correlated completely with ABO‐related haplotypes previously determined by Southern blot. Conclusion: The three Amerindian tribes represent a homogeneous (ABO blood group) population, except for the G542A mutation. The presence of this mutation in all other populations examined suggests that it originated before the migration of man into America.
New Murine Monoclonal Antibodies Directed against Glycophorins C and D, Have Anti‐Ge2 Specificity
Authors: Janvier D. , Veaux S. , Benbunan M.
Published: December 2004

Background and Objectives: The Gerbich blood group system comprises three high‐incidence antigens which have been localized on glycophorins C (GPC) and D (GPD) molecules. Murine monoclonal antibodies (mAbs) directed against Ge4 and Ge3 epitopes have already been obtained. We describe two murine mAbs (Z151.17 and J6) with Ge2 specificity. Materials and Methods: The mAbs were investigated by serological methods and immunoblotting, with common, Ge‐negative and enzyme‐modified RBCs. Results: Both mAbs behaved like anti‐Ge2 in serological tests, but bound to GPC and GPD on immunoblots. Z151.17 did not recognize the GPCYus or GPCGe variants, whereas J6 revealed a 25‐kD component from Ge:‐2,‐3,4 membranes, which is identical to the GPCGe molecule. Conclusion: These two mAbs are directed to slightly different epitopes. The Z151.17 and J6 epitopes are common to GPC and GPD, thus distinct from the Ge2 antigen defined by human alloantibodies.
Dra (Cromer‐Related Blood Group Antigen)‐Incompatible Renal Transplantation
Authors: Nakache Richard , Levene Cyril , Sela Ruth , Kaufman Suzana , Shapira Zaki
Published: December 2004

Background and objectives: Cromer system antigens, a series of blood group antigens of very high frequency, are not considered to be clinically significant in transfusion. In renal transplantation only the ABO blood group antigens are considered essential. The Drori blood group antigen is present in serum and has been found to reside on the renal tubular basement membrane and Bowman's capsule. The effect of anti‐Dra on the renal parenchyma has not been evaluated. Materials and methods: A unique case of renal transplantation of an incompatible Dr(a+) kidney to a Dr(a−) patient with anti‐Dra in her serum is presented. Results: Graft function was immediately good. The titer of anti‐Dra remained unchanged following transplantation. Conclusion: The successful outcome of a case of a Drori (Dra)‐incompatible kidney transplantation confirmed the lack of clinical significance of the anti‐Dra relating to transplantation.
α2,3‐Specific Desialylation of Human Red Cells: Effect on the Autoantigens of the Pr, Sa and Sia‐I1, ‐b1, ‐Ib1 Series
Authors: Roelcke D. , Hack H. , Kreft H. , Gross H. J.
Published: December 2004

Background and objectives: Pr1,2,3, PrM, Sa and Sia‐11, ‐b1, ‐1b1 are sialic acid (NeuNAc)‐dependent antigens recognized by human cold agglutinins. Pr and Sa antigens are the O‐glycans of glycophorins containing α2,3NeuNAc (to galactose) and/or α2,6NeuNAc (to galactosamine). The antigens of the Sia‐11, ‐b1, ‐1b1 complex are gangliosides that may carry α2,3NeuNAc (to galactose) and/or α2,8NeuNAc (to NeuNAc). We studied the NeuNAc groups involved in the antigens. Materials and methods: From 74 sera with cold agglutinins against NeuNAc‐dependent antigens, anti‐T‐free preparations were made and tested against human red cells, treated with an α2,3‐specific recombinant sialidase. Results: Most (51/62) Pr antigens use α2,3NeuNAc, some (8/62) use α2,6NeuNAc and a few (3/62) use both sialyl groups as immunodominant components on glycophorins. The immunodominant component of Sa and Sia‐11, ‐b1, ‐1b1 determinants was α2,3NeuNAc in all cases. Conclusion: The red cell target structures for cold agglutinins against NeuNAc‐dependent antigens have been identified. We propose a Pr nomenclature to reflect this. The binding of anti‐Pr to gangliosides may be the basis for anti‐Pr‐induced peripheral neuropathy.
Review of Counselling in a Transfusion Service: The London (UK) Experience
Authors: Miller R. , Hewitt P. E. , Warwick R. , Moore M. C. , Vincent B.
Published: December 2004

Donor (and recipient) counselling within the Transfusion Service in the UK has grown in volume and complexity over the last 10 years. The addition of new tests for donated blood and the growth of bone marrow transplantation have increased the demands on counselling staff. New initiatives, such as the HCV look‐back programme, have required an extension of the skills and knowledge of staff involved in counselling.
How Some Attitudes, Beliefs and Motivations of Spanish Blood Donors Evolve over Time
Authors: Fernández‐Montoya A. , López‐Berrio A. , Luna del Castillo J. D.
Published: December 2004

Objectives: To determine whether the attitudes, beliefs and motivations of blood donors evolve over time. Materials and Methods: Using questionnaires a 7‐year longitudinal study of a cohort of 126 donors was performed to gather their sociodemographic characteristics and their attitudes, beliefs and motivations relating to blood donation. Results: Changes were observed in a large number of beliefs and attitudes, with a reduction in fear about donation and in the need for rewards and recognition, and an increase in comfort during donation and in attitudes of duty and solidarity. However, the motivations of 65% of the sample did not change. Conclusion: With the passage of time, donors' attitudes and beliefs evolve in a way that is favourable to blood donation and their motivations remain stable.
Solid‐Phase Immunoassays for HCV Antibodies in Gamimune® N
Authors: Schreiner Clara , Musmanno Lisa , Kuenzi Mark , Torres Juan
Published: December 2004

Background and objectives: Occasional false‐positive results for the presence of HCV antibodies occur when testing pH 4 Gamimune® N immune globulin intravenous (human) (IGIV) using commercially available immunoassay kits. We investigated the causes. Materials and methods: EIA test results were compared with Nile red hydrophobic dye binding. Results: There was a strict correlation of reactive EIA results with a larger relative proportion of molecules in a low pH‐dependent hydrophobic conformation. Conversely, the greater the hydrophilic contribution to the population of IgG molecules (favored at neutral pH), the more nonreactive the EIA test results became for the same lots. An IGIV test preparation procedure consisting of pH neutralization followed by 48‐hour incubation at 37°C decreased positive test result outcomes in four different immunoassay formats by shifting the IgG population to a more hydrophilic conformation. Conclusion: The hydrophobic conformation of IgG favored at low pH appears to give rise to significant levels of nonspecific binding in solid phase immunoassays.
Epidemiological and Clinical Aspects of Hepatitis G Virus Infection in Blood Donors and Immunocompromised Recipients of HGV‐Contaminated Blood
Authors: Heuft H. G. , Berg T. , Schreier E. , Künkel U. , Tacke M. , Schwella N. , Hopf U. , Salama A. , Huhn D.
Published: December 2004

Background and objectives: The infectiousness and clinical relevance of the newly discovered blood‐borne Flaviviridae‐like agent, termed hepatitis G virus (HGV), are not well understood. Materials and methods: Twenty‐three transfusion recipients of two HGV‐affected long‐term blood donors were studied for HGV genome and antibodies to the putative envelope 2 glycoprotein (anti‐E2) of HGV. Nine recipients had nonhematological disorders and 14 suffered from severe hematological diseases and 7 of them received allogeneic bone marrow or blood stem cell transplantation. The molecular epidemiology of the observed HGV infection was studied by direct sequencing of parts of the 5′‐noncoding region, NS3, and NS5 region of HGV in the 2 long‐term donors and in their 6 recipients who became HGV RNA positive. Additionally, 549 individuals – homologous (n = 254) and autologous blood donors (n = 202), and medical staff (n = 89) – were investigated for the presence of HGV RNA. Results: HGV RNA in serum was found in 15 of the 23 (65%) transfusion reciients with known exposure of HGV‐contaminated blood. Seven of the remaining 8 recipients showed only an anti‐E2 response, indicating previous HGV infection with spontaneous clearance of the virus. In one recipient neither HGV RNA nor anti‐E2 could be detected. Molecular evidence for HGV transmission by the 2 donors was found in 3 of the 6 recipients studied. The alanine aminotransferase levels were not significantly different in the HGV RNA positive and negative recipients, and none of the 23 recipients developed posttransfusion hepatitis. Persistent HGV infection was observed especially in recipients with severe hematological disorders or in those in whome intensive immunosuppressive treatment was necessary. Of the 549 individuals studied, 10 (1.8%) were healthy carriers of HGV RNA. Conclusion: The persistence of transfusion‐acquired HGV infection is not associated with acute or chronic hepatitis, but may be influenced by the recipient's underlying disease.
Quality and Safety of Platelet Apheresis Concentrates Produced with a New Leukocyte Reduction System
Authors: Riggert J. , Humpe A. , Simson G. , Köhler M.
Published: December 2004

Objectives: Contaminating white blood cells (WBC) in apheresis platelet concentrates (PC) can cause a variety of adverse effects after platelet transfusion. To obtain PCs with low WBC contamination, a new leukoreduction system (LRS) utilizing ‘fluidized particle bed’ technology has recently been introduced. Methods: We prospectively examined the effect of LRS apheresis on the donor, the quality of the resulting PCs (n = 120), and the platelet increment in the corresponding recipients. Conventionally prepared apheresis PCs served as control group (n = 27). Platelet glycoproteins were examined by flow cytometry. Results: In LRS apheresis, we observed no serious adverse effects on the donors, but the postdonation absolute lymphocyte counts were reduced from 1,787±505/μl to 1,405±383/μl (p<0.001). Comparable results were seen in non‐LRS donors. The collection efficiency of the LRS procedures was 50.0±7.6%, resulting in a yield of 4.3±1.0 × 1011 platelets/PC. In flow cytometry, platelet glycoproteins in LRS PCs were not elevated: mean fluorescence of CD62 (6±4) or CD63 (9±3) in comparison with non‐LRS PCs (mean fluorescence of CD62: 7±4, CD63: 8±3). Median leukocyte contamination of the LRS PCs was 0.41 × 105 (range 0.07–8.5) WBCs/unit. In 43 recipients, the 24‐hour corrected count increments after transfusion of LRS PCs (12,530±8,761) were essentially the same as those of 20 recipients of non‐LRS PCs (13,133±9,812; p = 0.75). Conclusions: LRS apheresis appears to be a safe procedure, which produced effective PCs with few contaminating leukocytes. With new apheresis technology, filtration of PCs may become superfluous.
Evaluation of a Platelet Apheresis Technique for the Preparation of Leukocyte‐Reduced Platelet Concentrates
Authors: Zingsem Jürgen , Glaser Anke , Weisbach Volker , Zimmermann Robert , Neidhardt Bernd , van Waeg Geert , Eckstein Reinhold
Published: December 2004

Objectives: Reduction of the white blood cell (WBC) contamination in platelet concentrates (PC) protects patients from the immunological and infectious side effects of platelet transfusion caused by WBC. This can be done either by filtration of the PC or by improved apheresis techniques that yield WBC‐poor preparations. Methods: To evaluate an improved technique for platelet collection, we carried out 201 separations in 89 healthy cytapheresis donors using the new COBE Spectra leukoreduction system (LRS) and compared the results with those of standard dual‐needle separations obtained with the same cell separator. Results: A small but statistically significant difference was found in platelet collection efficiency in comparison with the standard non‐LRS software procedures (LRS: 52.6 vs. 56.3% for the reference). However, median WBC contamination was only 0.01 × 106 WBC per LRS product. This significant (p<0.0005) improvement corresponds to a 10‐fold reduction of WBC as compared with the standard dual‐needle technique. Conclusions: The COBE Spectra LRS system produced PCs with a platelet collection efficiency nearly equal to previous techniques and with a residual WBC content satisfying even the most stringent criteria for WBC‐depleted blood components. As this purity is achieved without important platelet loss, conventional fiber filtration no longer seems necessary in this kind of PC.
A Comparative Evaluation of the Sensitivity of Five Anti‐Hepatitis C Virus Immunoblot Assays
Authors: Couroucé A. M. , Noel L. , Barin F. , Elghouzzi M. H. , Lunel F. , North M. L. , Smilovici W.
Published: December 2004

Background and Objectives: In order to compare sensitivity, five anti‐hepatitis C virus (HCV) immunoblot assays were tested (Deciscan plus, Inno‐Lia III, Matrix, Murex Western blot and RIBA‐3). Materials and Methods: The test panel (50 samples for each assay) included 6 anti‐HCV‐negative samples and 44 samples from 36 HCV‐infected subjects. Results: There were minor differences in core reactivity among the assays. The smallest number of NS3‐reactive results occurred with the Murex and Matrix assays, and the smallest number of NS4 reactives with RIBA‐3 and Matrix. Among the 20 discrepant results for NS5 there was one clear false‐negative with Inno‐Lia. Only 28 of the 50 samples of the panel gave the same results in all the assays: 5 negatives and 23 positives. One of the 6 negative samples was indeterminate in 3 assays. Eighteen of the 21 other divergent results were interpreted as either indeterminate or positive, a common reactivity being exhibited by all 5 assays. The most important discrepancies occurred on 3 HCV‐RNA‐positive samples which came up negative in some assays: 2 samples with isolated NS3 reactivity were negative by Matrix and Murex Western blot, 1 of them being also negative by Inno‐Lia III; another sample was negative by RIBA‐3 and Matrix due to weak signals (<1) on core and NS3 proteins, which did not exceed 1+ with the other assays. Conclusions: With more uniform criteria for interpretation, the results would have been less divergent. Some assays should improve their sensitivity to the NS3 protein.
Evaluation of the Performance of Brazilian Blood Banks in Testing for Chagas' Disease
Authors: Saéz‐Alquézar Amadeo , Otani Márcia M. , Sabino Ester C. , Ribeiro‐dos‐Santos Gabriela , Salles Nanci , Chamone Dalton F.
Published: December 2004

Background and Objectives: Due to the low sensitivity and reproducibility of available tests, in 1989 it became mandatory for all Brazilian blood donors to be screened for Chagas' disease by at least two serological techniques. In 1994 the Brazilian Ministry of health launched a program to systematically evaluate the quality of serological screening for the detection of blood‐transmissible diseases as performed by public blood banks. Methods: A blind panel containing positive samples for blood‐transmissible disease was distributed to 57 major public blood banks in four sequential programs. Results: The ELISA test was chosen by the majority of the blood banks. There were 64 (3.7%) false‐negative results, 49 produced by banks using indirect hemagglutination. Since most blood banks screened with more than one test for Chagas, only 8 samples were actually missed, of which 3 were by banks using only one test. Conclusion: Our data show a clear improvement in performance of Brazilian blood banks testing for Chagas' disease.
Purification of α1 Proteinase Inhibitor from Human Plasma Fraction IV‐1 by Ion Exchange Chromatography
Authors: Chen Sharon X. , Hammond David J. , Lang John M. , Lebing Wytold R.
Published: December 2004

Background and Objectives: α‐proteinase inhibitor (PI) protects the lungs from proteolytic damage caused by elastase and can be used to treat congenital emphysema. We describe an improved method of purification of α1 PI from redissolved fraction IV‐1 paste. Materials and Methods: The process used dimethylaminoethy1 anion exchange chromatography, sulfopropyl cation exchange chromatography, virus inactivation by dry heat, and tri‐n‐butyl‐phosphate/cholate treatment, followed by a second strong cation exchange chromatography. Optimizations of loading conditions for ion exchange chromatography at small scale (20–60 ml of suspension) are described. Virus inactivation was adjusted to provide the best yield of α1 PI consistent with effective inactivation. The process has been effectively scaled up. Results: The final product was approximately 90% pure by SDS‐PAGE, with a 60–70% yield from starting fraction IV‐I paste. The process has been characterized by methods including nonreduced SDS‐PAGE, α1 PI inhibition assay, and biuret protein assay. Conclusion: The method described is an effective way of preparing large quantities of α1 PI from fractionated plasma.
Different Alleles Cause an Imbalance in A2 and A2B Phenotypes of the ABO Blood Group
Authors: Ogasawara Kenichi , Yabe Ryuichi , Uchikawa Makoto , Bannai Makoto , Nakata Kenichi , Takenaka Michiko , Takahashi Yuji , Juji Takeo , Tokunaga Katsushi
Published: December 2004

Background and Objectives: In several populations, including the Japanese, the frequency of the A2B phenotype is significantly higher than expected based on the A2 phenotype frequency. To understand the genetic basis of this ‘excess’ of A2B, we examined ABO alleles in individuals with A2‐related phenotypes. Materials and Methods: ABO alleles were identified by means of polymerase chain reaction single‐strand conformation polymorphism (SSCP) and nucleotide sequence analyses. Results: The frequencies of A2‐related alleles (*A105, *A106, *A107, *A111 and *R101) were clearly different between the A2 and A2B phenotypes. In particular, a putative recombinant allele, *R101, was uncommon in the A2 but common in the A2B phenotype individuals. This allele was also detected in 4 of 401 (1%) unrelated A1 phenotype (AO genotype) individuals. Conclusion: *R101 is presumably expressed as phenotype A1 in *R101/*O heterozygous individuals, but as phenotype A2 in *R101/*B heterozygotes, thus giving rise to a high A2B phenotype frequency.
Serum Affinity Chromatography for the Detection of lgG Alloantibodies in a Patient with High‐Titer lgM Cold Agglutinins
Authors: Stahl D. , Kreft H. , Hack H. , Schraven B. , Roelcke D.
Published: December 2004

Background and Objectives: High‐titer cold autoagglutinins (CA) interfere with the detection of alloantibodies against red‐blood‐cell antigens. The diagnostic procedure is extremely difficult, especially under pressure of time. It was the purpose of the study to find out whether quantitative IgG purification from serum of plasma by affinity chromatography facilitates the detection of IgG alloantibodies in the presence of high‐titer IgM CA. Materials and Methods: Serum: IgM‐ϰ CA, 0°C titer = 65,000; five anti‐IgG reactive alloantibodies. Affinity chromatography: HiTrap protein G 1‐ml affinity column (Pharmacia Biotech, Uppsala, Sweden). Results: All IgG alloantibodies were detectable in the eluate after affinity chromatography without interference of IgM CA. Conclusion: IgG alloantibodies in the presence of high‐titer IgM CA can be easily and rapidly detected under routine conditions by quantitative IgG purification from serum or plasma by affinity chromatography.
Transfusion Practices in Primary Total Joint Replacements in Finland
Authors: Capraro Leena
Published: December 2004

Background and Objectives: A multicenter survey was carried out in 11 Finnish hospitals to determine the prevailing transfusion practices in orthopedic surgery. Materials and Methods: The records of 1,161 patients who had undergone primary total hip (THR) or knee replacement (TKR) were reviewed. Results: Allogeneic red cells (RBC) were administered to 92% of the THR patients and to 84% of the patients undergoing TKR. Significant interhospital variation in RBC transfusion, from 2.4 to 4.1 units per patient in THR and from 1.8 to 2.8 units in TKR, was detected after adjustment for clinical and laboratory findings. Conclusion: The use of RBCs in orthopedic surgery was generous compared with reports from other European countries and the US. Each hospital's individual transfusion policy seemed to be a strong determinant of allogeneic blood use.
Point Mutations and Deletion Responsible for the Bombay H null and the Reunion H weak Blood Groups
Authors: Fernandez‐Mateos Pilar , Cailleau Anne , Henry Stephen , Costache Marieta , Elmgren Anders , Svensson Lola , Larson Göran , Samuelsson Bo E. , Oriol Rafael , Mollicone Rosella
Published: December 2004

Objective: Definition of the molecular basis of the Reunion and the Bombay red cell and salivary H‐deficient phenotypes. Methods: Sequence and expression of FUT1 and FUT2 genes from H‐deficient individuals. Family segregation analysis of the mutations responsible for the fucosyltransferase defects of H, secretor and Lewis systems. Results: The Indian red cell H null Bombay phenotype depends on a new mutation of the FUT1 gene. T725→G changing Leu242→Arg. Their salivary nonsecretor phenotype is secondary to a complete deletion of the FUT2 gene. The red cell H weak Reunion phenotype depends on another new mutation of FUT1, C349→T which induces a change of His117→Tyr. Their salivary nonsecretor phenotype is due to the known Caucasian inactivating mutation G428→A. Conclusion: Single prevalent FUT1 and FUT2 point mutations and a deletion are responsible for the Indian Bombay H null and the Reunion H weak phenotypes found on Reunion island. This is in contrast with other H‐deficient phenotypes where sporadic nonprevalent inactivating mutations are the rule.
Different Genotypes Causing Indiscernible Patterns of A Expression on A el Red Blood Cells as Visualized by Scanning Immunogold Electron Microscopy
Authors: Hansen Torjørn , Namork Ellen , Olsson Martin L. , Alan Chester M , Heier Hans Erik
Published: December 2004

Background and Objectives: The published sequence of the weak A subgroup Ael gene from Swedish individuals showed a G insertion in exon VII, causing a frameshift at ccdon 268 (the A01 gene has 353 codons). We wished to sequence exons VI and VII of two Norwegian Ael individuals and compare the expression of A substance on RBC from different A el individuals. Materials and Methods: Exon VI and VII were amplified by PCR, cloned in M13 and sequenced. A structure expression on Ael RBC was studied by the immunogold technique. Results: In contrast to the Swedish Ael individuals, the two Norwegians had consensus A01 sequences in exon VI and VII. However, the patterns of A expression were indiscernible from the Swedish cases as visualized by immunogold labeling in SEM. In both cases, a few (1–2%) RBC were very strongly labeled, some were weakly labeled and the majority (95%) were unlabeled. Conclusion: Although some Ael individuals have an inserted nucleotide in exon VII of the ABO gene, others have consensus A01 sequence in exon VI and VII. However, we could not find any differences in phenotype by immunogold labeling in SEM.
RHD Gene Polymorphisms among RhD‐Negative Chinese in Taiwan
Authors: Sun Chien‐Feng , Chou Cheng‐Sheng , Lai Nan‐Chang , Wang Wei‐Ting
Published: December 2004

Background and Objective: The rare occurrence of anti‐D‐associated hemolytic disease of the newborn among Chinese is attributable in part to the existence of the weak D phenotype Del among apparently RhD‐negative individuals. While exciting advances in the molecular genetics of the Rh blood group have been noted in recent years, the genomic structure of the Del phenotype has seldom been studied in the literature. We try to explore the genomic structure of the RhD gene among apparently Rh‐negative Chinese in Taiwan in this study. Methods: Genomic DNA from 230 samples of apparently RhD‐negative Chinese was studied using four polymerase chain reaction (PCR)‐based RhD typing methods. These PCR methods amplified RHD and RHCE genes at exons 4, 5, 7 and 10. All nuclieotides responsible for exofacial amino acid differences between RhD and RhCeEe peptides, including amino acids 169, 170, 172, 223, 226, 233, 238, 350, 353, and 354, were contained in these amplified DNA segments. Southern blot analysis using RHD cDNA fragments as probes was performed. Results: According to the serological study, 155 samples (67.4%) were genuinely RhD‐negative and 75 samples (32.6%) were of the Del phenotype. Successful amplifications for RHD sequences were possible in all 75 Del samples using four PCR methods. Apparently, all Del individuals carried an intact RHD gene. While 145 individuals of 155 genuinely Rh‐negative (63.0% of apparently RhD‐negative individuals) had total deletion of their RHD genes, 10 individuals (4.3% of apparently RhD‐negative individuals) were shown to have a preserved 3′ noncoding region of the RHD exon 10 and a gross deletion of RHD exons 4–10. Conclusions: Three classes of RhD‐negative polymorphisms among Chinese in Taiwan were observed. These included Del with grossly intact RHD and weak RhD expression, genuinely RhD‐negative with total deletion of the RHD gene. A molecular study is warranted to clarify the mechanism responsible for the weak RHD gene expression in Del individuals.
Characterization of a Macaque Anti‐Rh17‐Like Monoclonal Antibody
Authors: Blancher A. , Roubinet F. , Reid M.E. , Socha W.W. , Bailly P. , Bénard P.
Published: December 2004

In order to produce macaque monoclonal antibodies (mAbs) against human red blood cell (RBC) antigens, macaques were immunized with human and gorilla RBCs and their spleen lymphocytes were fused with man‐mouse heteromyeloma cells. One macaque‐mouse heterohybridoma produced a macaque IgGχ (Cyn2‐4D5) which agglutinated all human RBCs but not rare human variants Dc–, D––, and Rhnull. Thus, Cyn2‐4D5 exhibited RH17‐like reactivity. The specificity of Cyn2‐4D5 for RHCE‐encoded polypeptides was confirmed by specific immunoprecipitation of RhcE and RhCe polypeptides from human RBCs and the absence of immunoprecipitation of the RhD polypeptides extracted from D–RBCs. This study demonstrates that it is possible to produce macaque mAbs against human RBC blood group antigens.
The ELO Blood Group Polymorphism Is Located in the Putative First Extracellular Loop of Human Erythrocyte Band 3
Authors: Zelinski Teresa , Punter Fiona , McManus Kirk , Coghlan Gail
Published: December 2004

Background and Objectives: The low incidence red cell antigen ELO (700.51) has been excluded from 13 of 23 established blood group systems. Information relative to the Diego system has not been reported. To investigate a possible association between ELO and the gene controlling Diego blood group polymorphisms, we undertook molecular studies of AE1 (anion exchanger 1 gene). Materials and Methods: DNA from a family segregating for ELO and from the original ELO proposita was amplified by PCR using intronic primers flanking exons 11‐20 of AE1. Subsequently, single‐strand conformational polymorphism (SSCP) analysis and DNA sequencing were conducted. Results: We have observed an exon 12 SSCP in all ELO+ individuals tested. This SSCP is the result of a C→T mutation in exon 12 of AE1 which leads to an Arg432→ Trp amino acid substitution in the putative first extracellular loop of band 3 and thereby accounts for the Diego blood group system polymorphism known as ELO. Conclusions: In light of these findings, the International Society of Blood Transfusion Working Party on Terminology for Red Cell Surface Antigens has designated ELO as a member of the Diego blood group system (010008 or DI18) and rendered the previous numerical designation (700.51) obsolete.
A Simple Strategy to Look Back on Posttransfusion Hepatitis B in a Multitransfused Patient
Authors: Jongerius J. M. , van der Poel C. L. , van Leeuwen C. F.
Published: December 2004

Background and Objectives: In January 1996, a case of hepatitis B virus (HBV) seroconversion in a multitransfused patient was reported to the blood bank. From March through October 1995, the patient had received 23 units of red cells and 30 units of pooled platelet concentrates, encompassing an exposure to a total of 200 whole blood donations. Materials and Methods: In order to trace hepatitis B surface antigen (HBsAg)‐negative but HBV‐infectious blood donation(s), we tested samples of the donors obtained ≥3 months after the implicated donations for anti‐HBc (Corezyme EIA, Abbott). From 172/200 donors, archived samples of subsequent donations were available for this purpose. The remaining 28 donors were reinvited to the blood bank to obtain an additional blood sample for anti‐HBc testing. Results: 1/200 follow‐up donor samples was anti‐HBc‐positive. Retrospective testing of the implicated HBsAg‐negative blood donation of this donor revealed anti‐HBc‐negative and HBV‐DNA‐positive results. The patient was transfused with the platelets of the HBV‐infectious donation. On looking back, the other blood products prepared from this HBV‐infectious donation caused posttransfusion HBV infection (PT‐HBV) in 2 additional patients. Conclusion: Anti‐HBc testing on mainly archived follow‐up samples of 200 donors implicated in PT‐HBV was a rapid, simple, cost‐effective and donor friendly method to identify an infectious but HBsAg‐negative, anti‐HBc‐negative and HBV‐DNA PCR‐positive blood donation. Routine anti‐HBc screening would not have prevented this HBV transmission.
Unsatisfactory Detection of an in vivo Haemolytic Anti‐Vel by the Gel Test
Authors: Neppert J. , Bartz L. , Clasen C.
Published: December 2004

Background and Objectives: A patient experienced a severe haemolytic transfusion reaction. Neither the haemolytic property nor the specificity of the causative antibody had been sufficiently recognised when performing a microcolumn gel test. Materials and Methods: Subsequent to the transfusion reaction, the serological property and specificity of the causative antibody were analysed. Tube and gel test methods were compared, as were various reagent red cell specimens and their constituents. Results: A haemolytic anti‐Vel was detected in the tube test. In contrast, the particular commercial gel test kit used did not reveal the haemolytic property or specificity of the antibody. Our experiments suggest that this was apparently due to the presence of EDTA in the low ionic strength saline solution of the test kit. Conclusion: In rare cases life‐threatening haemolytic activity of an irregular blood group antibody may be undetected by a commercial microcolumn gel test kit in which EDTA is a constituent.