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Autoimmune hemolytic anemia is an uncommon disease that can be challenging to manage both for the emergency department physician and the general practitioner. The diagnosis is based on specific biological changes and on a positive direct Coombs test. Depending on the severity of the anemia and its clinical impact, an urgent blood transfusion can be required. However, ABO blood group typing and antibody screening may be impaired by autoantibodies. In case of vital need, a transfusion of ABO, Rh D and, if possible, C, c, E, e and Kell antigen matched red cells can be performed, before the complete achievement of the pre-transfusion testing. Further management includes the introduction of immunosuppression and the treatment of a possible underlying disease. Early contact with the hematologist, is strongly recommended.L’anémie hémolytique autoimmune (AHAI) est une pathologie peu fréquente, dont la prise en charge peut représenter un défi pour l’urgentiste et le généraliste. Le diagnostic est posé sur la base des modifications spécifiques du bilan biologique et sur la positivité du test de Coombs direct. Selon la sévérité de l’anémie, une transfusion sanguine en urgence peut s’avérer nécessaire. Les tests prétransfusionnels peuvent être perturbés par la présence des autoanticorps. En cas d’urgence vitale, on transfusera en compatibilité ABO, Rh D et en essayant de respecter les antigènes C, c, E, e et Kell, sans attendre les tests prétransfusionnels complets. La suite de la prise en charge s’articulera autour de l’introduction de l’immunosuppression ainsi que sur la recherche d’une maladie sous-jacente. L’appel précoce à l’hématologue est de rigueur.

Describe the management of Acquired Immune Haemolytic Anaemia (AIHA) and correlate with the current guidelines published in 2009. The secondary objective was to calculate the positive predictive value of the Direct Antiglobulin Test (DAT) for the diagnosis of AIHA.A retrospective and monocentric study was performed from 2010 to 2015 based on positive DATs, identified in the French Blood Agency database or in medical files. All patients managed for initial diagnosis or relapse of AIHA were included, excluding neoplasia.Six hundred and twenty-three patients had a positive DAT, 42 had non-neoplastic AIHA. Thirty-nine patients were included, 32 had warm antibodies, 5 had a negative DAT and 2 had cold antibodies. No cause was found for 46% (17/37) of the warm antibody and negative DATs AIHAs. Autoimmune disease was found in 11 cases (30%), infection in 4 cases (11%). The etiologic investigations were consistent with the guidelines in 49% of cases. Corticosteroids were first prescribed, as recommended. Second-line treatments were rituximab in 9 cases, splenectomy in 4 cases and azathioprine in 3 cases. The management of cold antibody AIHA complied with the guidelines. The positive predictive value of DATs in hospitalized population was of 14% (85/610).AIHA guidelines seem insufficiently applied in our center.

Evaluate the anti-erythrocyte and anti-HLA immunization rates in hemoglobinopathies.Cross-sectional study (October 2009-March 2010) on 83 patients followed for hemoglobinopathies. The irregular antibodies research is realized by two techniques: indirect Coombs and enzymatic technique on gel cards. The search for anti-HLA class I antibodies is done by complement dependent lymphocytotoxicity.The mean age was 30 years (14-64 years), the sex ratio M/F is 0.84. Our series included 42 cases of sickle cell disease (29 homozygous sickle cell anemia and 13 sickle-thalassemia) and 41 cases of thalassemia syndromes (26 major and 15 intermediate). The anti-erythrocyte alloimmunization rate is 10.84% without difference between thalassemia syndromes and sickle cell disease. The autoimmunization rate (22.89%) is higher in thalassemia syndromes (41.46%) than in the sickle cell disease (7.14%) (P0.001). The anti-HLA immunization rate is 31.6% without difference between thalassemia syndromes and sickle cell disease. The young age, transfusion at a young age and the total number of transfusions are the factors that increase the risk of anti-erythrocyte autoimmunization. No clinicobiological parameter does influence the anti-erythrocyte and anti-HLA alloimmunization. There is no significant association between anti-erythrocyte and anti-HLA immunization.The erythrocyte and anti-HLA anti-immunization rates are high in our series. Preventive strategy is needed to ensure optimal blood safety.

Red cell transfusion is one of the main treatments in sickle cell disease. However there are potential risks of blood transfusions. In order to propose strategies to improve blood safety in sickle cell disease in Mali, we conducted a prospective study of 133 patients with sickle cell anemia recruited at the sickle cell disease research and control center of Bamako, November 2010 to October 2011. The study aimed to determine the prevalence of human immunodeficiency virus (HIV), hepatitis B virus (HBV), and hepatitis C virus (HCV) infections by serum screening and the frequency of red cell alloimmunization before and after blood transfusion. The diagnosis of sickle cell syndrome was made by HPLC, the detection of markers of viral infection was performed by ELISA, and the diagnosis of alloimmunization was conducted by the Indirect Coombs test. Prevalence of viral infections observed at the time of enrolment of patients in the study was 1%, 3% and 1% respectively for HIV, HBV and HCV. Three cases of seroconversion after blood transfusion were detected, including one for HIV, one for HBV and one another for HCV in sickle cell anemia patients. All these patients had received blood from occasional donors. The red cell alloimmunization was observed in 4.4% of patients. All antibodies belonged to Rh system only. Blood transfusion safety in sickle cell anemia patients in Mali should be improved by the introduction of at least the technique for detecting the viral genome in the panel of screening tests and a policy of transfusions of blood units only from regular blood donors.

Molecular testing in red blood cell immuno-haematology is used extensively since the last 5 past years because of the technical developments and the possibility to use commercial kits. But these analyses have a high cost and rely on dedicated laboratories. Therefore, serology remains a reference technique for many laboratories in France, and in the world. Molecular analyses are used to resolve problems that cannot be resolved by serology. In this review, we will detail the main indications of molecular analysis, in donors and recipients, taking into account the technical tools that we can use, and the knowledge that we have about blood groups and their different variants. In this context, we have to remember that a molecular analysis has to be performed only if there is a benefice for the patient in the transfusion or the obstetrical setting.

The pattern of autoimmune hemolytic anemia has changed significantly these last 15 years. With regard to the diagnosis strategy, the use of gel filtration technique to perform the direct antiglobulin test (DAT) has decreased the number of autoimmune haemolytic anemias with negative tests results. In recent years, autoimmune haemolytic anemia increased in patients receiving purine nucleoside analogues, blood transfusions, solid organ transplantation or hematopoietic stem cells transplantation. These difficult autoimmune haemolytic anemia cases need to use new kinds of treatments. With regard to the treatment, very little progress was made this latter 50 years. The discovery of the efficacy of anti-CD20 antibody in this disease represents a breakthrough. Nowdays, the second-line treatment includes rituximab or splenectomy. Sometimes, the anti-CD20 treatment could be proposed in first-line but some clinical trials are needed.

To evaluate the efficiency of blood tests (blood group, direct antiglobulin test) to assess severe hyperbilirubinemia in full-term newborns, delivered from mothers with rhesus negative or O group and to determine clinical and biological factors that may improve the prediction characteristics of this blood test.We included all the full-term newborns, delivered from mothers with rhesus negative or O group, in a tertiary maternity ward, in 2005, from January6th to December31st.One thousand and ninety-two children were included. Newborns of A, B or AB group delivered from a mother 0 were at increased risk of presenting severe hyperbilirubinemia (OR=2.35 [1.22-4.52]). The negative predictive value was 96%. Yet, the determination of the Coombs test does not increase NPV.Systematic performance of blood test for newborns delivered from mother with O group does increase the ability to predict severe hyperbilirubinemia in a newborn infant. Direct antiglobulin test systematic performance remains questionable.

Autoimmune hemolytic anemia (AIHA) is diagnosed in the presence of anemia, usually macrocytic and of variable intensity, reticulocytosis, and a positive direct and/or indirect antiglobulin test, after ruling out other types of hemolytic anemia. A positive direct antiglobulin test alone is not sufficient to diagnose AIHA and may be positive in many patients without anemia or negative in some patients with AIHA. AIHA may be classified into two major categories according to the optimal temperature of antibody activity: warm-reacting autoantibodies (usually IgG) optimal around 37 degrees C and cold-reacting autoantibodies, optimal at 4 degrees C (usually IgM). This classification guides the selection of tests and treatment. AIHA is widely reported to be associated with a variety of other diseases, although these associations are often fortuitous. A minimal set of useful investigations is appropriate since AIHA may be secondary to viral infections, lymphoid malignancies, or autoimmune disorders such as lupus. Transfusion should remain rare in AHAI, but close contact with the transfusion service is necessary if it is to succeed. As for many autoimmune and/or systemic diseases, numerous types of treatment have been proposed but have not been validated in controlled multicenter studies. These are necessary to improve the management of these rare disorders.

Confirmation of autoimmune hemolytic anaemia usually relies on the detection of erythrocyte membrane-bound autoantibodies using a direct antiglobulin test. In the rare case of IgA autoantibodies-mediated autoimmune hemolytic anemia, the direct antiglobulin test can be negative, because routinely used polyspecific direct antiglobulin test reagents contain only anti-IgG and anticomplement antibodies.We report the case of a 41-year-old woman presenting a severe autoimmune hemolytic anaemia caused by the presence of warm autoantibodies of IgA type that revealed a chronic hepatitis C virus infection.A negative direct antiglobulin test does not completely rule out the diagnosis of autoimmune hemolytic anaemia especially in the rare case of IgA mediated immune hemolysis. The diagnosis strategy of autoimmune hemolytic anaemia associated with negative direct antiglobulin test and the potential links between autoimmune hemolytic anaemia and HCV are discussed.

Drug-induced immune hemolytic anemia is a rare cause of hemolytic anemia.A 68-year-old male patient developed an acute intravascular hemolysis with acute renal failure. Common causes of hemolysis were ruled out and the patient rapidly improved. An immune mechanism was confirmed by the positivity of the direct antiglobulin test with anti-IgA in the presence of ambroxol only, one of the drug the patient had received during 6 days before hospitalization.To our knowledge, this is the first report of ambroxol-induced immune hemolytic anemia. This case also underlined that the direct antiglobulin test should also be performed with anti-IgA to rule out any false negative.

The publication of the Circular DGS/DHO/Afssaps N(o) 2003-582 of the December 15 2003 relating to the administration of blood and blood components, means that an update of statutory requirements is needed. This article presents the overall process from the writing of prescriptions to the care and monitoring of the transfused patient. An update and a critical analysis of regulations currently in force are presented for each step as well as a description of the main adverse events.

Cross-matching between the serum of a patient and the red blood cells to be transfused is most important for the prevention of hemolytic transfusion reactions in allo-immunized or new-born patients found positive with direct antiglobulin test. Cross-matching is a time-consuming and complex laboratory test. In order to obtain valid results, it is necessary to abide by some technical rules detailed in this article. The choice of the blood units to be cross-matched depends on the patient's clinical story and on the specificity of anti-erythrocyte antibodies present in the serum. The identification and the management of most frequent difficulties met by using the cross-match technique are discussed hereby.

In a transfusional or foeto-maternal context, hemolysis by incompatibility due to anti-erythrocyte antibodies (regular or irregular) remains the most frequent and most serious immunological risk in the receiver. In order to prevent this risk, a number of actions must be taken, such as the realization of the immunohematologic analyses for which the methodological practices have been legislated because of their serious clinical consequences. Several elements play a role in the reliability of the analyses and their results: the selection of the reagents and their validation in the routine technique used; the validation of reception; the controls involved in secondary preparations (e.g., blood cells reagent); and the daily internal controls. All this requires the choice of adapted controls and the management of possible anomalies.

Autoimmune hemolytic anemias (AIHA) are characterized by hyperhemolysis associated with the presence of the immunoglobulins IgG, IgM or IgA on the red cell membrane. These immunoglobulins react as auto-antibodies against the red cell auto-antigens of the patient. The diagnosis is supported by clinical and biological signs of hemolysis, and by the identification of the auto-antibodies using the direct antiglobulin test (DAT). Here we report 14 cases of patients who showed the clinical and biological profile of AIHA, but who gave a negative DAT. We therefore tried to determine the presence of IgA on the red cell membrane with a method more sensitive than the DAT: the gel test using anti-IgA. With such a gel test, we demonstrated that there were IgA auto-antibodies on the red cell membrane in the 14 cases, therefore confirming the diagnosis of AIHA. We discuss the interest of performing a gel test with anti-IgA in each case where AIHA is suspected, but in which a negative DAT has been observed.

The detection of irregular antibodies is usually performed with serum by the indirect antiglobulin test (IAT) with a polyspecific antiglobulin reagent. In a first study in 1996, we compared the results obtained with 3,264 blood samples of patients drawn with or without anticoagulant: no significant difference was observed among the 240 allo-antibodies detected and identified. In this paper we report the comparison of the results obtained by IAT on column of filtration with two kinds of reagents: polyspecific and anti-IgG antibodies. Respectively 2,927 (76 contained an antibody), and 643 (161 contained an antibody) sera of patients were tested with methods ID-Diamed and Ortho-Biovue. Titrations of 153 other antibodies were also performed with the two reagents. Results showed no significant difference using either polyspecific reagent or the anti-IgG antibodies. This study proves that it is possible to perform screening of irregular antibodies on uncoagulated blood samples. This possibility allows automation as blood typing and screening of irregular antibodies can be carried out with the same sample.

The French Blood Transfusion Society working group Immunohaematology and the French Blood Transfusion Center of Lille performed two quality control exercises-96-01 and 97-02-in order to evaluate identification performances of alloantibodies associated with an autoantibody recognizing a high frequency antigen. Concerning control 96-01, 83 (75%) of the 110 blood transfusion centers participating at this exercise sent results. The alloantibody screening was correct for 78 of them (94%). Sixty-one (78%) blood transfusion centers correctly identified the specificities (anti-RH1 + anti-FY1). Concerning control 97-02, 82 (94%) of the 87 blood transfusion centers participating in this exercise sent results. The alloantibody screening was correct for 69 (88%) of them. Fifty-three blood transfusion centers (77%) correctly identified the specificities (anti-RH3 + anti-FY1). These exercises allowed us to confirm the main procedures used in routine for national scale testing. The analysis of the results has underlined the importance of these tests for assessing the quality of these examinations, and highlighted the means to be carried out in order to improve them.

The Société Française de Transfusion Sanguine and the Centre National de Référence pour les Groupes Sanguins performed a quality control to evaluate the performances of two serological tests: the Direct Antiglobulin Test (DAT) and the Elution test. Among the 110 Blood Transfusion Centers participating in this control, 80 (73%) returned a result. Of these, 68 results were correct for the DAT (85%; positive for type IgG) and 31 results were correct for the elution (39%; anti-FY1). This control gave the opportunity to confirm the main procedures used in routine testing on a national scale. The analysis of the results underlines the importance of the choice of a standardized technique for these tests. Such controls are useful to appreciate the quality of the routine tests and to find the means to improve them.

Coombs test and flow cytometry have been compared in terms of specificity and sensibility in a population of hospitalized patients, for whom a Coombs test had been required. The Coombs test seems more sensible than flow cytometry to detect red cell-bound IgG. For a given patient and over the time, flow cytometry seems better correlated with the severity of haemolysis if erythrocytes are strongly sensitized by IgG. The results of flow cytometric analysis, in percentage of sensitized erythrocytes, do not allow to define a sensibilization threshold for haemolysis prediction. Flow cytometric analysis would be more sensible than Coombs test in the detection of red cell-bound C3d, but these cases are not associated with autoimmune haemolytic anemia. Direct Coombs test should remain the diagnostic test of autoimmune haemolytic anemia.

Mycoplasma pneumoniae (MP) infection is associated with the emergence of anti-I cold agglutinins of IgM isotype, rarely complicated by hemolytic anemia. The mechanism by which the autoimmune disease occurs is still considerably controversed. We describe here a case of MP infection complicated by severe autoimmune hemolytic anemia with monoclonal anti-I IgM. IgM anti-I antibodies were purified by adsorption-elution on OI+ red cells. The red blood cell antibodies were found to be tightly associated with IgG and anti-MP specificity probably as immune complexes. The significance of these results is discussed.

The use of red blood cells coated with complement fractions using low ionic strength saline conditions is part of the evaluation of anti-complement antibodies, specially monoclonal antibodies used for antiglobulin reagents. This work shows that such cells stored in liquid nitrogen allow satisfactory long-term studies except for the anti-C3g antibodies.

To assess immune responses to malaria-induced thrombocytopenia, an haematologic and immunologic study was performed on 25 patients with imported malaria upon admission and 8 days after treatment. Thrombocytopenia (150 x 10(9)/litre) was detected in 19 cases (P. falciparum: 11 cases, P. ovale: 6 cases, P. vivax: 2 cases). No laboratory evidence of disseminated intravascular coagulation impairment was found in any of the patients. Bone marrow examination performed in 9 cases showed no abnormality in the megakaryocyte series. Platelet count was independent of circulating parasite levels (r = 0.27) and inversely related to the number of antibody binding sites (ABS) on platelets (r = -0.6, p. less than 0.01). The indirect Coombs test (r = -0.54; p less than 0.01) and IgG and IgE levels (p less than 0.02) gave similar findings. A statistical correlation was observed between the level of circulating immune complex and the number of ABS (r = 0.525, p less than 0.01). Thus malaria-induced thrombocytopenia seems to mainly involve IgG type antiplatelet antibody activity. Although they may be implicated in the binding of antibodies to platelets, circulating immune complexes do not appear to mediate thrombocytopenia.

The authors present 2 cases of mycoplasmal pneumonia associated with severe haemolysis. Haemolytic anaemia is a well-known and habitually mild complication of the disease. It is related to a peak of cold agglutinins, which are antibodies that agglutinate red blood cells usually at low temperatures but sometimes at 37 degrees C when they are present in high concentrations. The pathogenesis of cold agglutinins is thought to involve the secretion of peroxides by Mycoplasma pneumoniae with alteration of ed cell antigens which become immunogenic.

A french woman delivered a third full-term male baby who had a strongly positive direct antiglobulin test. During the pregnancy and after the delivery, the woman had a negative irregular antibody screening test using standard red blood cell panels. The compatibility testing between the mother's serum and the father's red blood cells was strongly positive and the antibody was identified as an anti-Wra. The baby developed a mild hyperbilirubinemia and recovered without treatment. This child was probably responsible for his mother's immunization since the two previous children were Wra negative and the mother had no history of blood transfusion or abortion.