Rapid screening of platelet donors for Pl^A1 (HPA-1a) alloantigen using a solid-phase microplate immunoassay

Abstract

Pl A1 and Pl A2 are alternative platelet-specific alloantigens in the Pl A1 system. Sensitization to Pl A1 underlies most cases of neonatal alloimmune thrombocytopenia (NAIT) and posttransfusion purpura (PTP) in white populations. A rapid and simple method for large-scale platelet phenotyping is desirable for identifying expectant mothers at risk of allosensitization and for identifying Pl A1 -negative donors when transfusions are indicated for treatment of NAIT or PTP. We investigated the effectiveness of a solid-phase microplate immunoassay for this purpose. Platelet-rich donor plasmas were tested using the Capture-P ® kit (Immucor, Norcross, GA). Platelet monolayers in microtiter wells were incubated with anti-Pl A1 , washed, and exposed to red blood cells (RBCs) precoated with anti-human IgG. Adherence of RBCs in a diffuse pattern across the well surface indicated the attachment of anti-Pl A1 to Pl A1 -positive platelets whereas sedimentation of unattached RBCs into a central pellet indicated the platelets were Pl A1 -negative. Of 520 donors, 15 (2.88%) tested Pl A1 -negative, which correlates well with the reported Pl A2,2 frequency in whites of 2.25 percent. Results were confirmed by DNA genotyping and/or immunoblotting. This screening technique permits phenotyping donors for Pl A1 alloantigen with minimal specialized equipment. Confirmatory testing for Pl A2 alloantigen can be reserved for donors that test negative for Pl A1 .