Diagnosis of Neonatal Alloimmune Thrombocytopenia: Determination of the Specificity of Antiplatelet Alloantibodies in the Maternal Blood Plasma Using a Molecular and Genetic Method

Background: Thrombocytopenia occurs in 1-5 % of newborns (platelet count < 150 × 109/L). Low platelet 50 109/L leads to the hemorrhagic syndrome, with one its causes being neonatal alloimmune thrombocytopenia resulting from incompatibility between mother and fetus human antigens (HPA) inherited father absent mother, which formation maternal antibodies. Anti-HPA-1a, anti-HPA-5b, anti-HPA-3a, anti-HPA-3b antibodies are clinically significant as they destroy fetal/neonatal platelets causing severe complications (intracranial hemorrhage 20 cases prenatal or postnatal death 10 cases). Adequate diagnosis is a key successful treatment approach, largely depends on cause. Objective: To determine nature specificity mother’s blood. Materials methods: We studied blood samples parents (21 pairs) Saint Petersburg, Russian Federation. used flow cytometry alloantibodies plasma after incubation paternal staining Goat F(ab’)2 Anti-Human IgG-FITC CD41-PE monoclonal Allosensitization index was calculated percentage IgG-positive cells number fixing anti-CD41 At value ≥ 15 %, antiplatelet were considered present sample. molecular detection system FluoVista analyzer (Inno-Train, Germany) for genetic testing allele-specific primers. Alleles genes encoding expression HPA-1, HPA-2, HPA-3, HPA-4, HPA-5, HPA-6, HPA-9, HPA-15 determined by real-time polymerase chain reaction using set HPA-FluoGene reagents Germany). Genomic DNA isolated DNA-sorb-B (AmpliSens, Federation). Results: found that 8 21 (38 %) mothers had against platelets. During genotyping mother/father pairs, incompatible combinations revealed: HPA-1b/HPA-1a 9 pairs (HPA-1a antigen platelets), 5 (55%) probable anti-HPA-1a; HPA-1a/HPA-1b 4 2 (50 having an anti-HPA-1b specificity. HPA-3a/HPA-3b observed (probably anti-HPA3b) 1 (25%). HPA-2a/HPA-2b, HPA-5a/HPA-5b, HPA-15a/HPA-15b, HPA-15b/HPA-15a incompatibilities detected (1 case each pairs), no mothers. The distributed follows: 62 anti-HPA-1a, 25% anti-HPA-1b, 13 anti-HPA-3a. Conclusions: confirmed immune cases.