Multicenter Study on Differential Human Neutrophil Antigen 2 Expression and Underlying Molecular Mechanisms.

Background: The human neutrophil antigen 2 (HNA-2), which is expressed on CD177, is undetectable in 3-5% of the normal population. Exposure of these HNA-2 individuals to HNA-2-positive cells can cause immunization and pro-duction of HNA-2 antibodies, which can induce immune neutropenia and transfusion-related acute lung injury. In HNA-2-positive individuals, neutrophils are divided into a CD177 and a CD177 subpopulation.

The association and functional relevance of genetic variation in low-to-medium-affinity Fc-gamma receptors with clinical platelet transfusion refractoriness.

Background: Inadequate responses to platelet transfusions (i.e., platelet transfusion refractoriness [PLT refractoriness]) are a serious problem.

Progress and development of platelet antibody detection.

Human platelet antibody (HPA) detection is necessary for the diagnosis and therapeutic decisions for refractoriness to platelet transfusions, post transfusion purpura and fetal and neonatal alloimmune thrombocytopenia. In the last four to five decades many new developments, both in knowledge and methods, have increased the quality of platelet serology. However, the quest for the optimal antibody detection method(s) encountered, sometimes unexpected, difficulties.

Fast and low-cost direct ELISA for high-throughput serological HPA-1a typing.

Background: Fetal and neonatal alloimmune thrombocytopenia (FNAIT) is caused by maternal alloantibodies against fetal human platelet antigens (HPAs), mostly caused by anti-HPA-1a. Population-based screening for FNAIT is still a topic of debate. Logistically and financially, the major challenge for implementation is the typing of pregnant women to recognize the 2% HPA-1a-negative women.

Plasma thrombopoietin levels as additional tool in clinical management of thrombocytopenic neonates.

Plasma thrombopoietin (Tpo) levels distinguish thrombocytopenia resulting from increased platelet destruction or decreased platelet production. We investigated whether measuring plasma Tpo levels in thrombocytopenic newborns is of diagnostic value to establish the underlying mechanism of thrombocytopenia.Tpo levels were measured with in-house developed ELISA in samples referred to our center because of thrombocytopenia noticed in the first 10 days of life.

Detection of platelet autoantibodies to identify immune thrombocytopenia: state of the art.

Immune Thrombocytopenia (ITP) is diagnosed by exclusion of other causes for thrombocytopenia. Reliable detection of platelet autoantibodies would support the clinical diagnosis of ITP and prevent misdiagnosis. We optimized our diagnostic algorithm for suspected ITP using the direct monoclonal antibody immobilization of platelet antigens assay (MAIPA), which evaluates the presence of platelet autoantibodies on the glycoproteins (GP) IIb/IIIa, Ib/IX and V bound on the patient platelets.

A multicenter validation of recombinant β3 integrin-coupled beads to detect human platelet antigen-1 alloantibodies in 498 cases of fetomaternal alloimmune thrombocytopenia.

Background: Fetomaternal alloimmune thrombocytopenia (FMAIT) is caused by human platelet (PLT) antigen (HPA) incompatibility. Beads coupled with recombinant β3 integrins, displaying the biallelic HPA-1 epitopes (rHPA-1), have been shown to detect HPA-1a alloantibodies implicated in FMAIT. This report describes a multicenter validation of the beads using the results of well-characterized samples to define the optimum parameters for analysis of a large cohort of 498 clinical samples.

Management and outcome of 35 cases with foetal/neonatal alloimmune neutropenia.

Aim: The aim of this study was to provide an overview of foetal/neonatal alloimmune neutropenia (FNAIN), together with advice on the clinical management.

Methods: Neutrophil serology in the Netherlands is centralised at Sanquin Diagnostic Services. We examined FNAIN cases between January 1, 1991, and July 1, 2013, to determine the number of cases diagnosed, the relationship with human neutrophil antigen (HNA) antibody, the clinical presentation and therapeutic interventions.

A new bead-based human platelet antigen antibodies detection assay versus the monoclonal antibody immobilization of platelet antigens assay.

Background: The performance of a newly developed Luminex bead-based platelet (PLT) antibody detection method (PAKLx) was compared with the monoclonal antibody immobilization of PLT antigens (MAIPA) assay and the LifeScreen Deluxe Luminex bead-based HLA Class I antibody detection method (LMX).

Study Design And Methods: Six sera containing anti-human PLT antigen (HPA)-1a (n=2), HPA-1b, HPA-2b, HPA-3a, or HPA-5b were tested in titration. A total of 194 sera, including HPA-1a, -1b, -2a, -2b, -3a, -5a, and -5b antibodies with or without HLA antibodies (n=63); glycoprotein (GP) IV antibodies (n=1); PLT autoantibodies (n=3); HLA antibodies (n=45); and samples with no PLT-reactive antibodies (n=82), were tested in both assays.

Neonatal alloimmune neutropenia due to immunoglobulin G antibodies against human neutrophil antigen-5a.

Background: Neonatal alloimmune-mediated neutropenia (NAIN) due to maternal alloantibodies directed against one of the human neutrophil antigens (HNAs) can cause severe infections. NAIN has been described as caused by antibodies against HNA-1a, -b, -c, -2a, -3a, or -4a, but not by antibodies against HNA-5a.

Results: Blood from a 3-week-old newborn and from his parents was sent to our laboratory because of suspicion of NAIN.

Alloantibodies against low-frequency human platelet antigens do not account for a significant proportion of cases of fetomaternal alloimmune thrombocytopenia: evidence from 1054 cases.

Background: Maternal alloantibodies against the five common human platelet antigen (HPA) systems (HPA-1 to -3, -5, and -15) are found in only 20% of cases referred for fetal and neonatal thrombocytopenia (FMAIT) investigations. The question asked was whether mismatches for the remaining 11 low-frequency HPAs (HPA-4 and -6bw to -17bw) might in part explain the remaining 80% of cases.

Study Design And Methods: A total of 1054 paternal DNA samples from referred FMAIT cases (among which 223 cases where antibodies against a common HPA were found) were genotyped for 11 low-frequency HPAs as well as a recently discovered polymorphism (ITGA2B-C2320T).

Acquired Glanzmann's thrombasthenia caused by glycoprotein IIb/IIIa autoantibodies of the immunoglobulin G1 (IgG1), IgG2 or IgG4 subclass: a study in six cases.

Background: Acquired Glanzmann's thrombasthenia (GT) is an uncommon bleeding disorder caused by glycoprotein (GP) IIb/IIIa-specific autoantibodies. Covering of the fibrinogen binding site of GPIIb/IIIa results in a moderate-to-severe bleeding tendency.

Materials And Methods: We performed a diagnostic evaluation and evaluated the underlying risk factors in six patients with a bleeding tendency caused by acquired GT.

Three novel beta3 domain-deletion peptides for the sensitive and specific detection of HPA-4 and six low frequency beta3-HPA antibodies.

Background: Antibodies against human platelet antigens (HPA) are clinically important in fetal-maternal alloimmune thrombocytopenia, refractoriness to platelet transfusions and post-transfusion purpura. Of the 16 HPAs, nine are located on the beta3 subunit of the alphaIIb beta3 integrin. Antibody detection is generally based on platelet-derived alphaIIb beta3 from HPA-genotyped donors.

Glycoprotein V: the predominant target antigen in gold-induced autoimmune thrombocytopenia.

Autoimmune thrombocytopenia is generally caused by autoantibodies against glycoprotein (GP) IIb-IIIa or GPIb-IX and occasionally against GPIa-IIa or GPV. By investigating 38 rheumatoid arthritis (RA) patients on gold therapy, 10 with profound thrombocytopenia and 28 nonthrombocytopenic controls, we showed that in all 10 patients with thrombocytopenia, the platelet autoantibodies preferentially targeted GPV but the presence of gold was not required for their reactivity. Elevated levels of platelet-associated IgG (PAIgG) were observed in 8 of the 10 patients in whom the tests were performed.

The diagnostic value of thrombopoietin level measurements in thrombocytopenia.

It has been reported that blood trombopoietin (TPO) levels can discriminate between thrombocytopenia due to increased platelet destruction and decreased platelet production. With our TPO ELISA and a glycocalicin ELISA we analysed a large group of patients in detail and could confirm and amplify the above notion in detail. TPO levels were determined in plasma from 178 clinically and serologically well-defined thrombocytopenic patients: 72 patients with idiopathic autoimmune thrombocytopenia (AITP), 29 patients with secondary AITP, 5 patients with amegakaryocytic thrombocytopenia and 72 patients who suffered from various diseases (46 in whom megakaryocyte deficiency was not and 26 in whom it was expected).

Max(a), a new low-frequency platelet-specific antigen localized on glycoprotein IIb, is associated with neonatal alloimmune thrombocytopenia.

We have identified a new platelet-specific alloantigen, Max(a), responsible for a typical case of neonatal alloimmune thrombocytopenic purpura. The maternal serum reacted strongly with paternal platelets in the platelet immunofluorescence test, whereas platelet alloantigen typing showed that no known human platelet antigen (HPA)-system was involved. In the monoclonal antibody (MoAb)-specific immobilization of platelet antigens (MAIPA) assay, the new antigen was located on the platelet membrane glycoprotein (GP) IIb-IIIa complex, but immunoprecipitation and immunoblot experiments to further localize the antigen failed.

Determination of human platelet antigen frequencies in the Dutch population by immunophenotyping and DNA (allele-specific restriction enzyme) analysis.

Platelets from 200 random Dutch blood donors were typed for the human platelet alloantigens HPA-1 to -5 recognized at present and for Naka. Naka is an epitope on glycoprotein IV, not expressed on the platelet of individuals with hereditary GP IV deficiency. Platelet immunofluorescence and monoclonal antibody-specific immobilization of platelet antigens (MAIPA) were applied for this purpose.

Protection against immune haemolytic disease of newborn infants by maternal monocyte-reactive IgG alloantibodies (anti-HLA-DR).

The extent to which maternal anti-Rh(D) antibodies support lysis of erythrocytes by monocytes in the antibody-dependent cell-mediated cytotoxicity (ADCC) assay is closely correlated with the severity of Rh(D) haemolytic disease of the newborn infant (HDN). However, in some cases HDN is much milder than would be predicted from the ADCC value. We postulated that maternal ADCC-blocking alloantibodies against paternal antigens on monocytes can protect these infants against severe haemolysis.

Complement is not activated in ABO-haemolytic disease of the newborn.

We studied the lysis in vitro of group A red cells by IgG anti-A. IgG anti-A, which strongly lysed A red cells from adults, did not lyse A red cells from cord blood, if fresh cord serum from a child with blood group AB was used as a source of complement. In cases of haemolytic disease of the newborn due to A-O or B-O antagonism with a positive direct antiglobulin test with anti-IgG serum, the red cells did not react with anti-complement sera.