Human platelet antigen 1a epitopes are dependent on the cation-regulated conformation of integrin α IIbβ 3 (GPIIb/IIIa)

David L. Allen*, Sofia Abrahamsson, Michael F. Murphy, David J. Roberts

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

11 Citations (Scopus)


Background: The HPA-1a (Leu 33) polymorphism of platelet integrin αIIbβ3 is the target of alloantibodies in 70-80% cases of neonatal alloimmune thrombocytopenia (NAIT) in Caucasians and reliable detection of these antibodies is essential for appropriate clinical management. However, the ability to detect such antibodies is highly variable between laboratories and, in a number of clinical cases where there is a HPA-1 genotype mismatch between mother and neonate, HPA-1a antibodies are undetectable. Furthermore, some studies have not shown a consistent relationship between maternal anti-HPA-1a level and neonatal platelet count. Since the integrity and conformation of the αIIbβ3 complex are dependent on divalent cations, we investigated whether HPA-1a epitope integrity and/or conformation might be affected by the presence of the cation chelator EDTA in patient samples or in assay buffers, thus providing a possible explanation for the variable sensitivity of current assays. Principle findings: Exposure of the αIIbβ3 complex to EDTA resulted in reduced reactivity of three anti-HPA-1a mAbs (B2, 19-7 and 23-15). More significantly, cation chelation adversely affected detection of polyclonal anti-HPA-1a, not only in the platelet immunofluorescence assay, where alloantibody binding was reduced compared to control platelets (mean MFI reduction 44.5%, range 17.3-69.7%, n= 4), but also in the commonly used monoclonal antibody specific immobilisation of platelet antigens assay (MAIPA) where both alloantibody and monoclonal capture antibody binding were reduced (mean OD reduction 82.8%, range 68.3-96.6%, n= 9). Conclusions: These data show that HPA-1a antibodies recognise epitopes on αIIbβ3 that are sensitive to EDTA treatment and that cation chelation grossly reduces the sensitivity of the MAIPA assay by diminishing not only HPA-1a alloantibody binding but also 'capture' monoclonal antibody binding. These findings may, in part, explain the current variability in antibody measurement and will guide the development of more sensitive tests for anti-integrin antibodies in NAIT and other conditions.

Original languageEnglish
Pages (from-to)166-175
Number of pages10
JournalJournal of Immunological Methods
Issue number1-2
Publication statusPublished - 31 Jan 2012
Externally publishedYes

Bibliographical note

Funding Information:
This project was approved by the Research and Development Committee of NHS Blood and Transplant (NHSBT) (Research Project No: PG99/13E/RD12) and by Mid and South Buckinghamshire Ethics Committee (LREC No: 06/Q1607/40). All donors gave their written informed consent and were identified to the researchers only by means of a unique ‘participant number’.


  • Alloantibody
  • Human platelet antigen
  • Integrin
  • NAIT
  • αIIbβ3


Dive into the research topics of 'Human platelet antigen 1a epitopes are dependent on the cation-regulated conformation of integrin α IIbβ 3 (GPIIb/IIIa)'. Together they form a unique fingerprint.

Cite this