Immunohaematological Tests Research Articles

Haemoglobin levels in autoimmune haemolytic anaemias at diagnosis: relationship with immunoproteins on red blood cells

Previous investigations of the relationship between characteristics of immunoproteins on red blood cells (RBCs) and the occurrence of autoimmune haemolysis yielded divergent results. Here, we studied these characteristics in autoimmune haemolytic anaemias (AIHAs) to determine their relationship with the degree of anaemia at diagnosis. We studied at diagnosis 52 cases of warm AIHA with positive direct antiglobulin test. Immunohaematological testing and determination of immunoglobulin class, complement, and immunoglobulin G (IgG) were performed using gel technology (GCT). Median haemoglobin (Hb) levels significantly differed between cases with IgG1 only or negative for IgG subclasses (7.4g/dl), those with IgG3 or IgG1+IgG3 (6.5g/dl), and those with multiple immunoglobulins (5g/dl). Logistic regression indicated that IgG3 detection was the only variable significantly related to the occurrence of RBC transfusion in AIHA (odds ratio 4.05, 95% CI 1.1-14.7). In our study, the type of immunoprotein(s) on the RBC surface was associated with different Hb levels at AIHA diagnosis. IgG3 and multiple immunoglobulins were associated with lower Hb levels; IgG3 was also associated with a higher percentage of patient transfusions in the first week after diagnosis. Thus, qualitative differences in these immunoproteins may lead to deeper and more prolonged anaemia levels, influencing the need for RBC transfusion.

Polyethylene glycol antiglobulin test (PEG-AGT)

Abstract Polyethylene glycol (PEG) was described in 1987 as a new technique for immunohematology testing. The original paper described its use in detection and identification of weakly reactive antibodies. PEG is used as an additive to enhance reactivity and to reduce incubation time when testing for unexpected antibodies. PEG can be used as an alternative to low-ionic-strength saline and whenever weak reactions are encountered. Immunohematology 2014;30:158–160.

Red Blood Cell Alloimmunization in Sickle Cell Disease Patients in Tanzania.

Alloimmunization is a recognized complication of red blood cell (RBC) transfusion and causes delayed hemolytic transfusion reactions and provides problems sourcing compatible blood for future transfusions. The objective of this study was to determine the frequency of RBC alloimmunization in SCD patients in Tanzania where pretransfusion screening for alloantibodies is not practiced. In a cross-sectional study, SCD patients at Muhimbili Hospital Sickle Cell Clinic, Dar es Salaam, Tanzania, were investigated. The demographic characteristics and transfusion history were recorded. Blood samples were drawn from consenting, previously transfused patients and RBC alloimmunization was demonstrated using immunohematologic techniques. There were 365 patients (median age, 16 years; 55.3% female) and they had received a median of 2 transfusion episodes. Fifteen patients (4.1%) possessed RBC alloantibodies. A total of 61 alloantibodies was found; 16 (26.2%) and 11 (18.0%), were directed against Kell and Rh blood group antigens, respectively. The rate of RBC alloimmunization in Tanzanian SCD patients was 4.1%. The low transfusion load may explain this immunization frequency. Nevertheless, our study confirms the significance of RBC alloimmunization as a complication in Tanzanian SCD patients. Therefore, there is need to improve immunohematologic testing in Tanzania so that RBC alloimmunization and its consequences may be prevented.

A New Method For Electronic Crossmatch: ABO /Rh Blood Group Confirmation and Antibody Screening Concomitantly With Serologic Crossmatch

Introduction Altough electronic crossmatch (E-XM) is cost effective and reduce laboratory workload, practical difficulties of determining 2 consecutive ABO/Rh in patients, requirement for the verification of the donor ABO group, concerns for errors in patient identification/specimen labeling and lack of reimbursement for performing ABO confirmation testing on a second separately collected specimen could avoid its widespread implementation in some countries. Method Using gel santrifuge method, ABO/Rh grouping and immuno-hematologic tests are done by a fully automatic system and recorded in blood bank information system. For patients and donors who do not have a previous blood group and antibody screen in the system, study steps are: 1. ABO/Rh group is performed with the 1st specimen and recorded 2. Donor blood group and antibody screen are done in the same gel card. 3. A second specimen is required at the time of transfusion request for regular antiglobulin crossmatch and antibody screen. ABO confirmation of patient and donor is done concomitantly in the same card with XM. 4. When 2 or more units are requested, first unit is selected by serologic XM, subsequent units are selected by E-XM from the available donor pool. Donor pool for electronic selection is fed by serologically compatible but undelivered units. Computer system is capable of questioning availability for E-XM eventhough serologic XM is not performed and could show the missing parameters needed for E-XM. Results After 6 months of validation process, between 1.01.2012- 31.07.2013, 36.7% of erythrocytes were reserved by E-XM using this method (table 1). Evaluation of the efficacy of the XM process revealed that 37.5% of the serologically confirmed units were undelivered and those supported the pool for E-XM (table 2). Conclusions 1. This method implements a double specimen system for ABO/Rh verification to ensure the transfusion security and guarantees the selection of at least 1 unit serologically. 2. Reduces work load and cost effective even in centers who are currently using serologic XM. Disclosures: No relevant conflicts of interest to declare.

Hyperhaemolysis syndrome responsive to splenectomy in a patient with δβ-thalassaemia: a discussion on underlying mechanisms.

Hyperhaemolysis syndrome responsive to splenectomy in a patient with δβ-thalassaemia: a discussion on underlying mechanisms.

Evaluation of Erytra® fully automated analyser for Routine Use in Transfusion Laboratory

To evaluate efficiency and performance of Erytra® analyser in comparison to the reference platform of the NBT immunohaematology laboratory (Bio-Rad ID-System). Moving to automation or semi-automation is a major focus of transfusion centres. The Erytra® (Grifols) is a new fully automated walk-away analyser with high volume processing capacity for pre-transfusion testing, designed to be used with the unique 8-column DG Gel® cards. A total of 2201 immunohaematological tests (1041 ABO/D grouping, 1041 antibody screening, 51 antibody identification, 45 newborns (ABO/D and DAT) and 23 crossmatches were performed on 1160 donor/patient whole blood samples. Erytra®'s performance was assessed by means of a stress test replicating the routine work of a hospital laboratory. Concordant results between the Erytra® and the reference method were obtained in 2195 (99·73 %) of the tests. There were only three discrepancies out of 6246 reactions (0·05%) in ABO/D grouping, all in the reverse group which did not mislead to group identification. Of the 1041 samples screened for antibody presence, Erytra® detected all the relevant antibodies [9 not detected weak prophylactic anti-D were determined to be clinically nonsignificant (<0·1 IU mL(-1) )] while Bio-Rad ID-System missed one anti-e and one anti-Jk(a) . Concordance for D grouping, crossmatching and newborns was 100%. Results of the simulated stress test exercise highlighted the capacity of Erytra® for absorbing into 4 h workloads equivalent to 24 h of routine. Grifols' Erytra® analyser showed reliable high sensitivity, velocity and capacity to cope with high workload in the immunohaematology laboratory routine.

Autoimmune Hemolytic Anemia With Gel-Based Immunohematology Tests

We used gel centrifugation tests (GCTs) to analyze the relationship between the diagnosis and immunohematology tests used for autoimmune hemolytic anemia (AIHA). The study included 588 samples positive for the direct antiglobulin test (DAT). Of these, 52 were from patients diagnosed with AIHA. Immunoglobulin (Ig) class, IgG1, IgG3, and complement were measured. DAT strength had the strongest correlation with AIHA diagnosis (odds ratio [OR], 23), followed by anti-IgG titer 300 (OR, 8.4), anti-IgG titer 1,000 (OR, 10.5), and C3d agglutination strength (OR, 1.7). Decision tree analysis revealed that DAT strength and anti-IgG titer higher than 100 were the best predictors of AIHA. Multidimensional scanning analysis found a high grade of similarity among DAT strength, anti-IgG titer, and IgG strength in the AIHA samples. This observation was not detected in DAT-positive samples from patients without AIHA. DAT strength remained the best diagnostic indicator for AIHA and had the strongest association with AIHA compared with other commercially available immunohematology tests. The other tests, despite good correlation with AIHA diagnosis, did not add useful information.

Severe warm autoimmune haemolytic anaemia due to anti-Jk(a) autoantibody associated with Parvovirus B19 infection in a child.

Severe warm autoimmune haemolytic anaemia due to anti-Jk(a) autoantibody associated with Parvovirus B19 infection in a child.

Pour une organisation transfusionnelle sécuritaire aux risques et coûts maîtrisés : l’expérience APHM–EFSAM

IntroductionBlood transfusion safety depends on strict compliance with each step of a process beginning with the order for labile blood products and related immunohematologic testing and ending with administration and follow-up of the receiver. This process is governed by stringent regulatory texts and guidelines. Despite precautions, processing errors are still reported. Analysis of incident reports shows that the most common cause involves patient identification and that most errors occur at two levels, i.e. the entry of patient information and management of multiple regulatory crosschecks and record-keeping using different systems. MethodThe purpose of this report is to describe the collaborative approach implemented by the Établissement français du Sang Alpes-Méditerranée (EFSAM) and the Assistance publique des Hôpitaux de Marseille (APHM) to secure the blood transfusion process and protect interfaces while simplifying and facilitating exchanges. ResultsClose cooperation has had a threefold impact with simplification of administration, improvement of experience feedback, and better management of test ordering. The organization implemented between the two institutions has minimized document redundancy and interfaces between immunohematologic testing and delivery. Collaboration based on experience feedback has improved the level of quality and cost control. ConclusionIn the domain of blood transfusion safety, the threshold of 10−5 has been reached with regard to the risk of ABO errors in the distribution concentrated red cells (CRC). In addition, this collaborative organization has created further opportunity for improvement by deploying new methods to identify simplification measures and by controlling demand and usage.

The importance of immunohematology testing in the neonatal period

In order to diagnose hemolytic disease of the newborn it is necessary to determine ABO/Rh blood group, direct antiglobulin test and indirect antiglobulin test in newborns as well as ABO/Rh and indirect antiglobulin test in their mothers due to a possible incompatibility between the blood types of the mother and her baby. The study was aimed at reviewing and analyzing the results of screening of the newborns and pregnant women on the territory of South Backa District during 2011, as well as at estimating the necessity to give blood transfusion to the newborns in the same period. Data obtained from the information system and protocol of the Institute for Blood Transfusion of Vojvodina were used in a retrospective analysis of screening of newborns and pregnant women. Blood samples taken from 3313 newborns were tested and the following distribution of the ABO blood types was recorded: O in 45.6%, A in 35%, B in 15.9%, AB in 3.5%; 78.9%, newborns were found to be D-positive, 21.1% were D-negative. The direct antiglobulin test was positive in 165 (4.9%) cases, it was weakly positive +/- in 85 (51.5%), positive 1+ in 45 (27.3%), positive 2+ in 17 (10.3%), positive 3+ in 12 (7.3%), positive 4+ in 6 (3.6%). In all cases, the monospecific direct antiglobulin test was positive with IgG antibodies. Of four newborns with positive direct antiglobulin test who required exchange blood transfusion, two had RhD incompatibility and two had ABO incompatibility. Blood samples of 3429 (100%) women were tested and 36 (1.05%) were actively immunized: 23 (0.67%) to RhD antigen, 8 (0.23%) to blood group antigens other than ABH and Rh, specificity could not be determined in 5 (0.15%). Anti-D antibodies were found in 19 (0.55%) pregnant women, in two (0.06%) cases newborns required exchange blood transfusion. Testing neonates and pregnant women contributes to the detection of blood type incompatibility between the mother and her child; it provides an opportunity for clinicians to implement the adequate prevention of RhD alloimmunization as well as to make timely diagnosis and to introduce treatment of hemolytic disease in newborns.

Providing compatible blood to patients with alloantibodies in ne Slovenia

Background: Erythrocyte alloantibodies often occur, especially in the case of polytransfusion and in pregnant women. Fast diagnosis and a compatible transfusion of the blood component are important to prevent an early or delayed post transfusion haemolytic reaction. We have analysed our efficiency to provide red cell units for these patients in our hospital and in the general hospitals of Ptuj and Murska Sobota. Methods: We analysed retrospectively which examinations had been done in sensitized patients, how successful we have been in finding appropriate blood for erythrocyte transfusion. We also analysed the proposed algorithm of the immunohaematological testing in our Centre. Results: At the Centre for transfusion medicine–CTM Maribor, we have provided compatible transfusions to 67 patients in the last year, for which we made 6215 tests to provide appropriate blood components. We were looking for compatibility between the serum of the patient and erythrocytes of the blood donor by a technique in which the antibodies showed the highest reactivity. In patients with antibodies of the Rh-system (D, E, e, C, c), Kell, Kidd, Duffy and MNS, we always provided antigen negative component, and in others a compatible component (negative result of cross match). Sometimes the appropriate blood was found based on already defined antigens in blood donors by means of the information system. If the appropriate component was found at the Unit for Transfusion Medicine in Ptuj or Murska Sobota the crossmatch was done there and the interpretation was done at the CTM by means of the teleconsultation system. Conclusions: In most cases we provided blood components successfully and relatively quickly, while in some cases, other transfusion centres were helping us out, especially the Blood Transfusion Centre Ljubljana and the Transfusion centre Celje.

ABO Hemolytic Disease Leading to Hyperbilirubinemia in Term Newborns: Value of Immunohematological Tests

Introduction ABO hemolytic disease of the newborn is most common cause of neonatal jaundice; previous studies have shown a poor correlation between serologic tests and clinical course in affected infants. The aim of this study was to identify the value of direct and indirect Coombs’ immunohematological tests, to identify the incidence of hemolytic disease in newborns (ABO-HDN). Methods : This two-year retrospective study of 1800 jaundiced term-neonates studied 355 cases due to ABO incompatibility divided into two groups: with and without hemolytic disease. Relation of laboratory parameters and immunohematological tests to severity of disease were studied. We did not analyze the maternal antibody titer or elusion test, and this was a limitation of the study. Results: In this study, 355 (19.7%) of all jaundiced newborn infants were ABO incompatibles; 98 (27.6%) of the newborns who had ABO incompatibility showed ABO-HDN (5.4% of total icteric patients). The positive direct antiglobulin (direct Coombs’ test) and indirect antiglobulin (indirect Coombs’ test) were diagnostic in 18.2% and 25.5% respectively in affected infants. The overall prevalence of immunohematological tests associated with ABO-HDN was 43.7%. There was significant correlation between positive antiglobulin tests and severity of jaundice (P=0.000); also there was a significant difference between indirect and direct Coombs’ test and severity of jaundice (P= 0.002). Conclusion:The antiglobulin tests, namely the indirect Coombs’ (IC) test and direct Coombs’ (DC) test are very useful to detect the newborns liable to serious jaundice.

Neonatal sepsis - Role of certain immunohematologic tests in rapid diagnosis

Background and Objective : Many neonates with sepsis die worldwide due to lack of early diagnosis and prompt treatment. This study was carried out to analyze various immunological and hematological parameters singly and in combination to formulate a guideline for the rapid diagnosis of neonatal sepsis. Materials and Methods : In a prospective study, 62 patients having clinical suspicion of neonatal sepsis were evaluated with a battery of investigations. Neonates admitted for other causes and without clinical suspicion of sepsis were selected as controls ( n = 40). The tests included blood culture, hemoglobin (Hb) level, total and differential blood count, absolute neutrophil count (ANC), immature to total neutrophil count (I/T ratio), micro-erythrocyte sedimentation rate (m-ESR), C-reactive protein (CRP), platelet count, serum immunoglobulin M (IgM), and plasma fibrinogen level. Patients were divided into proven cases (positive blood culture) and probable cases (blood culture negative). Results : Positive blood culture was seen in 38 cases (61.3%). Raised m-ESR (>8 mm in 1 st h) was seen in 63.2% of proven and 66.7% of probable cases. An I/T ratio of ≥0.2 was seen in 63.2% and 58.3% of proven and probable cases. Morphological changes were detected in 68.4% of proved and 91.7% of probable cases. Positive CRP test (≥6 mg/L) was found in 84.2% of proven and 100% of probable cases. Raised serum IgM, leukopenia, and neutropenia were seen in minority of patients (11-37%). Raised fibrinogen (>400 mg/L) was seen in the patients as well as in controls. Conclusions : Four useful tests were m-ESR, I/T ratio, morphological changes in neutrophils, and CRP; and were statistically significant ( P < 0.05). Most sensitive test was CRP (84%) and m-ESR was most specific test (94%). Combination of three or more of these four tests were highly specific (95-100%).

Neonatal sepsis: Role of a battery of immunohematological tests in early diagnosis.

Background and Objective:Worldwide, many neonates with sepsis die due to lack of early diagnosis. In this study we attempt to analyze the value of various immunological and hematological parameters, singly and in combination, for the diagnosis of neonatal sepsis, with the aim being to formulate guidelines for the early diagnosis of the condition.Materials and Methods:In this prospective study, 62 patients having clinical suspicion of neonatal sepsis were evaluated with a battery of investigations. Neonates admitted for other causes and without clinical suspicion of sepsis were selected as controls (n=40). The tests included blood culture, hemoglobin level, total and differential blood count, absolute neutrophil count, ratio of immature to total neutrophil count (I/T ratio), micro-erythrocyte sedimentation rate (m-ESR), C-reactive protein (CRP), platelet count, serum IgM level, and plasma fibrinogen level. Patients were divided into proven cases (positive blood culture) and probable cases (negative blood culture).Results:Positive blood culture was seen in 38 cases (61.3%). Raised m-ESR (>8 mm in the first hour) was seen in 63.2% of proven and 66.7% of probable cases. I/T ratio of ≥0.2 was seen in 63.2% and 58.3% of proven and probable cases, respectively. Morphological changes in neutrophils were detected in 68.4% of proven cases and 91.7% of probable cases. Positive CRP test (≥6 mg/l) was found in 84.2% of proven cases and 100% of probable cases. Raised serum IgM, leucopenia, and neutropenia were seen in a small number of patients (11%–37%). Raised fibrinogen level (>400 mg/l) was seen in patients as well as in controls.Conclusions:The four useful tests that we identified were m-ESR, I/T ratio, morphological changes in neutrophils, and CRP; and role of these tests in early diagnosis of neonatal sepis were statistically significant (P<.05). The most sensitive test was CRP (84%) and the most specific test was m-ESR (94%). A combination of three or all of these four tests was highly specific (95%–100%).

Automation in Immunohematology

There have been rapid technological advances in blood banking in South Asian region over the past decade with an increasing emphasis on quality and safety of blood products. The conventional test tube technique has given way to newer techniques such as column agglutination technique, solid phase red cell adherence assay, and erythrocyte-magnetized technique. These new technologies are adaptable to automation and major manufacturers in this field have come up with semi and fully automated equipments for immunohematology tests in the blood bank. Automation improves the objectivity and reproducibility of tests. It reduces human errors in patient identification and transcription errors. Documentation and traceability of tests, reagents and processes and archiving of results is another major advantage of automation. Shifting from manual methods to automation is a major undertaking for any transfusion service to provide quality patient care with lesser turnaround time for their ever increasing workload. This article discusses the various issues involved in the process.

Mechanical and electrical properties of red blood cells using optical tweezers

Optical tweezers are a very sensitive tool, based on photon momentum transfer, forindividual, cell by cell, manipulation and measurements, which can be applied to obtainimportant properties of erythrocytes for clinical and research purposes. Mechanical andelectrical properties of erythrocytes are critical parameters for stored cells in transfusioncenters, immunohematological tests performed in transfusional routines and in blooddiseases. In this work, we showed methods, based on optical tweezers, to study red bloodcells and applied them to measure apparent overall elasticity, apparent membrane viscosity,zeta potential, thickness of the double layer of electrical charges and adhesion in red bloodcells.

Participation in proficiency programs and promotion of quality in transfusion services of Minas Gerais

ObjectiveThis study aimed at identifying associations between the participation of transfusion services in immunohematology external quality control programs and their accuracy in immunohematology testing and adaptation to technical and legal operational procedures.MethodsFrom 2007 to 2009, a cross-sectional study was conducted in 219 transfusion services of the State of Minas Gerais who participated in this investigation by responding to a questionnaire and conducting a proficiency test comprising: ABO and RhD phenotyping, irregular RBC antibody screening and cross-matching. Frequencies and bivariate analysis followed by binary logistic regression were used for statistical analysis.ResultsTransfusion services who participated in external quality control programs (32.4%) and those that did not (67.6%) obtained worrying error percentages in proficiency tests which may significantly increase blood transfusion risks. Shortfalls related to the establishment of protocols, standards and internal quality control were also significant. On comparing the two groups, transfusion services that participated in these programs had a 2.35 times higher chance of correct results in the proficiency panel testing, a 3.16 higher chance of having transfusional records and a 2.81 higher chance of performing preventive maintenance of equipment.ConclusionThe study showed that independent factors associated to participation in external quality control programs suggest that more investment in internal quality control procedures is necessary and that more attention should be paid to current legislation.

Red blood cell alloimmunization in sickle cell disease patients in Uganda

Blood transfusion is an integral part in the management of sickle cell disease (SCD) patients. Alloimmunization is a recognized complication of red blood cell (RBC) transfusions with consequences including delayed hemolytic transfusion reactions and difficulties in getting compatible blood for future transfusions. The objective of this study was to determine the frequency of RBC alloimmunization in SCD patients in Uganda where pretransfusion screening for alloantibodies is not practiced. In a cross-sectional study, SCD patients at Mulago Hospital Sickle Cell Clinic, Kampala, Uganda, were investigated. The demographic characteristics and transfusion history were recorded. Blood samples were drawn from consenting, previously transfused patients and RBC alloimmunization was demonstrated using immunohematologic techniques. There were 428 patients (median age, 12 years; female/male ratio, 1.0) and they had received a median of 3 units in a median of three transfusion episodes. Twenty-six patients (6.1%) possessed RBC alloantibodies and 21 (80.7%) of them had received up to 10 transfusions. A total of 30 alloantibodies was found; 20 (66.7%) and 5 (16.6%) belonged to Rh and MNS blood groups, respectively. Five of the alloimmunized patients had multiple antibodies. The rate of RBC alloimmunization in Ugandan SCD patients was 6.1%. The homogeneity between donors and SCD patients plus the low transfusion load may explain this immunization frequency. Nevertheless, our study confirms the significance of RBC alloimmunization as a complication in Ugandan SCD patients. Therefore, there is need to improve immunohematologic testing in Uganda so that RBC alloimmunization and its consequences may be prevented.

Detection, Identification and Titration of Anti- C (RJ I4) Allo-Antibody in a Multi-Transfused HHA Patient Referred to the Department of Transfusion Medicine of BSMMU

It is a Report of a case of transfusion induced alloimmunization of anti-c (Rh-4) in multi transfused HHA.Following top-up transfusion DHTR resu lted in this multi transfused HHA. Patients was referred to the transfusion medicine department of BSMMU to detect the cause/s of DHTR from the Hematology department of Bogra MCH. High titrated (1: 256) anti-c alloantibody were detected, identified in this patient by the standardized sensitive method of immunohematological testing at the department of transfusion medicine, BSMMU. When incompatible RCCs arc transfused the amount of antibody in recipient's serum may be too low to effect red cell destruction or even to be not detected by sensitive compatibility tests. However Transfusion may provoke as anamnestic immune response so that a few days after transfusion a rapid increased in antibody concentration develops and rapid destruction of transfused red cells occur. Hemoglobinuira is not uncommon in DHTRs. and causes HDN During TOP-UP transfusion to each and every HHAs genotypically matched antigen negative RCC to be practiced to avoid DHTR. Unlike AIHA it is possible to identify and detect alloantibody responsible for the DHTR by meticulous immunohematological testing and processing.TAJ 2009; 22(1): 272-275

Loss of red cell ABH Antigens and Hypermethylation of the ABO Gene Promoter region is frequent in Myeloid Malignancies.

Abstract 3151Poster Board III-88 BackgroundLoss of red blood cell (RBC) antigens may occur in solid tumors and in hematological diseases; however the mechanisms involved in these changes are not fully understood. Aberrant DNA hypermethylation is thought to be involved in acute myeloid leukemia (AML) as well as in myelodysplastic syndromes (MDS), and the methylation of cytosines residues in the dinucleotide CpG may account for the expression patterns of the ABO genes. In this study we investigated loss of RBC ABH antigens and the possible role of DNA methylation in the ABO promoter gene in patients with myeloid diseases. MethodsForty-three patients were included in our study (MDS=16, chronic myeloid leukemia (CML)=12, AML=4, polycythemia vera (PV)=3, essential thrombocythemia (ET)=3, chronic myelomonocytic leukemia (CMML)=3, myelofibrosis (MF)=1 and chronic neutrophilic leukemia (CNL)=1). Forty-one patients were evaluated according to their ABO group using serological immunohematological tests (Diamed Inc., Brazil). ABH secreted antigens were investigated in saliva and a PCR-based ABO genotyping using restriction enzyme digestion (Alu and Kpn) was performed to confirm the ABO blood type. In addition, the expression of the A, B and H antigens was analyzed by flow cytometry in 26 patients (median age=63 yrs; 6M/17F) and 81 healthy controls (median age=33 yrs; 38M/43F). Methylation of CpG islands was investigated in 45 bone marrow samples using methylation specific PCR (MSP) technique with methylated and unmethylated primer sets for region from -200 to +26 sequence of the ABO gene. ResultsThe investigation of the secreted antigens in saliva and the genotyping studies confirmed the results of the ABO serological tests in 40 patients, but we found that the RBCs of one patient were not agglutinated by anti-B while his genotype result was compatible with BO indicating loss of the B antigen in his RBCs. Overall, loss of A, B or H antigen was detected by flow cytometry in 11/26 (42%) patients. Hypermethylation of the ABO promoter gene was detected in 51% (23/45) of the analyzed bone marrow samples. Among patients with hypermethylation in the ABO promoter gene, 44% had loss of A or B antigens confirmed by serological or flow cytometric tests compared with 28% in the group of patients with unmethylated ABO promoter gene (p=0.632). ConclusionsEpigenetic changes including methylation of cytosine residues are recognized as major contributors to gene silencing, disease progression and worse outcome in several cancer patients. Our data showed that the hypermethylation of the ABO gene is frequent in patients with myeloid malignancies corresponding to the pathogenesis already described for AML and MDS. However, not all patients showing loss of RBCs antigens had ABO methylation evidence, suggesting that other mechanisms may take place. Patients with myeloid malignancies often need blood transfusion support and loss of RBCs antigens can lead to changing in ABO blood group increasing the risk of serious blood transfusion reactions. The relatively high rate (42%) of loss of ABH antigens found in this study demonstrated that these patients have to be carefully managed to avoid such severe transfusion reactions. (Grants supported by CNPq, 478814/2006-2). DisclosuresNo relevant conflicts of interest to declare.