Francine Décary

Alloimmunization to the PlA1 platelet antigen : results of a prospective study

The natural history of alloimmunization to the PlA1 platelet antigen is uncertain. We followed 50 PlA1‐negative pregnant women during pregnancy and for 6 months post‐partum in order to determine this natural history. The cohort of PlA1‐negative women was obtained by PlA1 typing 5000 women. Three PlA1‐negative women formed anti‐PlA1 antibodies during this prospective study, two in pregnancy and one in the immediate post‐partum period. All three PlA1 antibody producers were HLA‐DR3 positive, a histocompatibility phenotype that is strongly associated with alloimmunization to the PlA1 antigen. One of the three infants delivered to these mothers was thrombocytopenic (platelet count 9 x 109/1). The remaining two infants had normal platelet counts at birth (160 and 174 x 109/1). The HLA‐A1, ‐B8, ‐DR3 and ‐DRw52 phenotype frequencies in the group of PlA1‐negative women who did not form PlA1 antibodies (n= 47) was similar to that found in their husbands, and that expected in a normal Caucasian population. From our data we estimate that alloimmunization to the PlA1 antigen occurs in approximately one out of every 1000 pregnancies in a Caucasian population. It is important to recognize that not all pregnancies in which a mother has formed PlA1 alloantibodies will result in the delivery of a thrombocytopenic infant. These findings are relevant to programs designed to either prevent alloimmunization to the PlA1 antigen (through passive administration of anti‐PlA1 immunoglobulin to at‐risk PlA1‐negative mothers), or to identify women at risk of delivery of thrombocytopenic infants (by antenatal screening to detect women alloimmunized to the PlA1 antigen).

Alloimmunization to Platelet antigen HPA-1a (PIA1) is strongly associated with both HLA-DRB3∗0101 and HLA-DQB1∗0201

Antibodies to the platelet HPA-1a antigen can elicit in the newborn a condition known as neonatal alloimmune thrombocytopenic purpura (NAITP). Previous studies based on RFLP analysis showed that 100% of HPA-1a-negative women who produced anti-HPA-1a antibodies (responders) were HLA-DRw52a (DRB3*0101). However, this specificity could also be found in some HPA-1a-negative women not producing anti-HPA-1a antibodies (nonresponders). We have analyzed in detail by PCR-SSOP the HLA-DR, -DQ, and -DP loci of 36 responders and 10 nonresponders. We found that while the allele DRB3*0101 was present in the vast majority of responders (91%), there were exceptions. Furthermore, the DQB1*0201 allele was found to be present in almost all responders (94%), but again was also found in nonresponders. The risk of alloimmunization to HPA-1a in an HPA-1b homozygous mother significantly increases with the presence of either allele, the odds ratio being 39.7 for DQB1*0201 and 24.9 for DRB3*0101. Sequencing of exon 2 of these two alleles from responders indicated no sequence difference when compared with the consensus sequences. This indicates that they do not represent variants when compared with the same alleles found in some nonresponders.

Evaluation of donor skin disinfection methods

BACKGROUND: Because most bacteria isolated from contaminated platelet concentrates are thought to originate from the donors skin, the efficacy of four methods of skin disinfection was compared.

Analysis of immunoglobulin class, IgG subclass and titre of HPA‐1a antibodies in alloimmunized mothers giving birth to babies with or without neonatal alloimmune thrombocytopenia

We analysed the titre and isotype composition of antibodies produced by mothers giving birth to babies with or without neonatal alloimmune thrombocytopenic purpura (NAITP) and patients with post‐transfusion purpura (PTP). All these individuals produced an antibody specific for the HPA‐1a allotype present on the platelet glycoprotein IIb‐IIIa (GPIIb‐IIIa). Sera from mothers who gave birth to thrombocytopenic babies (group 1, n = 36), non‐thrombo‐cytopenic babies (group 2, n = 4) or from PTP patients (group 3, n = 3) were tested by an indirect‐ELISA. Results indicated no evident differences in the isotype composition or titre of the antibodies from the three groups of sera. The antibody titre ranged from 1: 120 to 1: 3500. Antibodies with the IgGl subclass were present in all sera. Most sera contained IgGl alone (24/43 sera tested) or in combination with IgG3 (10/43). IgG2 was never present and only three sera showed intermediate reactivity with anti‐IgG4 MAb. Few sera (nine sera from groups 1 and 2) were weakly positive when tested with the anti‐IgM antibodies. These results suggest that neither the titre nor the isotype composition can be used to predict the severity or the occurrence of thrombocytopenia in newborns.

Elimination of heteroduplex artifacts when sequencing HLA genes amplified by polymerase chain reaction (PCR).

The polymerase chain reaction (PCR) can be used for DNA sequence analysis. When a unique sequence is amplified, the products generated are homogenous and can be sequenced directly. However, when multiple genes or alleles are amplified at the same time, such as in the case of HLA genes, the products generated are a heterogenous mixture that has to be cloned into bacteria before sequencing. In the latter case, variant sequences can arise during the PCR procedure as a result of in vitro recombination or of heteroduplex formation. In vitro recombination events result from end pairing of incompletely amplified sequences; synthesis is then completed and the resulting molecules amplified. A number of authors (Jansen and Ledley 1990; Lawlor et al. 1991; Meyerhans et al. 1990; Reynaud et al. 1991) have reported this phenomenon. However, the proportion of recombinant molecules to the total products of amplification is usually low (1-5%; Jansen and Ledley 1990; Meyerhans et al. 1990) and recombinants are relatively easily identified. Furthermore, the shorter the DNA template is, the lower is the risk of being incompletely synthesized by the Taq polymerase and of generating recombinants. Heteroduplex molecules are generated when fully synthesized complementary strands originating from distinct genes or alleles hybridize together. Jansen and Ledley (1990), when analyzing cDNA clones of the human methylmalonyl CoA mutase gene, estimated that 50 % of the final amplified products can be heteroduplexes. Actually the formation of heteroduplexes between PCR amplified homologous sequences has been used for the identification of heterozygotes, the detection of mutations

Nomenclature of Platelet-Specific Antigens

In order to avoid confusion, the Working Party on Platelet Serology decided to formulate a new nomenclature system for platelet-specific antigens.

Prenatal Management of Fetal Alloimmune Thrombocytopenia: Editorial

Alloimmune thrombocytopenia (AITP) of the fetus and newborn is a serious disease in which cerebral haemorrhage, which may occur as early as in the 18-20th week of pregnancy, may be fatal or lead to lasting damage to the brain. Optimal prenatal treatment of this disease is therefore essential. Since there did not seem to be a consensus of opinion about the optimal prenatal treatment, anumber of international experts were asked for their opinion. The answers to some of the questions confirm the lack of a general consensus.

Recognition of a Non-HLA-ABC Antigen Present on B and T Lymphocytes and Monocytes only Detectable with the Indirect Immunofluorescence Test

Abstract. The serum of a 39–year‐old male under long‐term platelet transfusion therapy for hypoplastic anemia with thrombocytopenia was investigated for the presence of leukocyte and platelet antibodies after the patient had received platelet concentrates from more than 700 random donors. The serological studies of his serum revealed: (1) the absence of plateletreactive antibodies; (2) the absence of agglutinating or cytotoxic antibodies against leukocytes; (3) the presence of at least two granulocyte‐specific antibodies, one with the specificity anti‐NA2 and the other with an undefined specificity, both only detectable by indirect immunofluorescence, and (4) the presence of cytotoxic‐negative fluorescence‐positive peripheral‐blood‐mononuclear‐cell‐reactive antibodies, not directed against HLA‐A, B or C antigens. The significance and implications of these findings are briefly discussed.

Alloimmunization to platelet antigen HPA-1a (Zwa): association with HLA-DRw52a is not 100%.

SIR, Neonatal alloimmune thrombocytopenic purpura (NAITP) is a disease of the new-born involving the destruction of the platelets of the fetus by maternal antibodies. In the Caucasian population, the incidence of NAITP is of about one in 2,000 births with an estimated morbidity and mortality of 15-30%. The majority of NAITP cases are associated with the HPA1 alloantigen system (designated previously as Zw or P1*), which consists of two allelic variants HPA-I a and

Iron and cardiac ischemia: a natural, quasi‐random experiment comparing eligible with disqualified blood donors (CME)

The theory that elevated iron stores can induce vascular injury and ischemia remains controversial. We conducted a cohort study of the effect of blood donation on the risk of coronary heart disease (CHD) by taking advantage of the quasi‐random exclusion of donors who obtained a falsely reactive test for a transmissible disease (TD) marker.