Inverted erythrocyte membranes demonstrate beta 2GPI-antiphospholipid antibody interactions and membrane crosslinking

Saartje Bloemen*, Xiao XuanWu, Katrien M. Devreese, Bas de Laat, Jacob H. Rand, Ljiljana V. Vasovic

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review


Introduction: The antiphospholipid syndrome (APS) is an acquired autoimmunc disorder predisposing patients to thrombosis or pregnancy complications. Since inverted erythrocyte membranes (iEMs) might provide a physiologically relevant source of anionic phospholipids, we studied the interactions of phospholipid-binding proteins and APS antibodies using iEMs. Materials & methods: iEMs were prepared from packed erythrocytes by hypotonic lysis. Phosphatidylserine (PS) exposure was confirmed by annexin A5 (A5) binding using fluorescence microscopy and flow cytometry. Binding of beta(2)-glycoprotein I (beta(2)GPI)-IgG immune complexes to iEMs was investigated with gel electrophoresis, western blot and flow cytometry. Functional involvement in coagulation was documental in the thrombin generation assay. Results: iEMs readily precipitated purified beta(2)GPI as well as beta(2)GPI from normal plasma and APS plasma. The plasma of APS patients provided higher levels of IgG binding to iEMs relative to healthy controls. Thrombin generation increased with increasing concentrations of iEMs, documenting that coagulation proteins bound to the exposed phospholipids. The LA effect was also distinguished in thrombin generation when comparing APS patients, as indicated by an increased lag time. Agglutination was observed after incubation with APS patient plasma and this was augmented by anti-human globulin. Conclusions: In conclusion, iEMs can provide a more physiological approach than phospholipid vesicle-based tests for investigating, APS and are more amenable to standardization than platelet membranes.
Original languageEnglish
Pages (from-to)89-94
JournalThrombosis Research
Publication statusPublished - Oct 2016


  • Antiphospholipici syndrome
  • Antiphospholipid antibody
  • Beta 2-glycoprotein I
  • Erythrocyte membrane
  • Immune complexes
  • Steric interactions

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