OC 23.2 - In vitro generated megakaryocytes for the detection of HPA alloantibodies in clinical samples

Sunday, June 25, 2023

15:00 – 15:15 ET

Room: 710B

Session Available As: On-Demand

Presenting Author(s)

Günalp Uzun, MD (he/him/his)

Physician
Center for Clinical Transfusion Medicine, University Hospital of Tübingen
Tübingen, Baden-Wurttemberg, Germany

Co-Author(s)

JL

Josip Lucic

Physician
Department of Transfusion Medicine, Cellular Therapeutics and Hemostaseology, Clinic for Anesthesiology LMU University Hospital Munich, Germany.
München, Bayern, Germany
FR

Flavianna Rigoni

MTA
Center for Clinical Transfusion Medicine
Tübingen, Baden-Wurttemberg, Germany
FL

Franziska Lyshy

MTA
Center for Clinical Transfusion Medicine
Tübingen, Baden-Wurttemberg, Germany
IM

Irene Marini

PhD
Center for Clinical Transfusion Medicine
Tübingen, Baden-Wurttemberg, Germany
SN

Stefanie Nowak-Harnau

Physician
Center for Clinical Transfusion Medicine
Tübingen, Baden-Wurttemberg, Germany
Karina Althaus, MD

Physician, Director of Platelet Immunology Lab.
Center for Clinical Transfusion Medicine, University Hospital of Tübingen
Tübingen, Baden-Wurttemberg, Germany
Ulrich Sachs, PhD

Physician
Institute for Clinical Immunology, Transfusion Medicine and Hemostasis, Justus-Liebig-University, Langhansstr. 7, 35392 Giessen, Germany.
Giessen, Hessen, Germany
Tamam Bakchoul, MD (he/him/his)

Medical Director
Center for Clinical Transfusion Medicine, University Hospital of Tübingen
Tübingen, Baden-Wurttemberg, Germany

Background: Detection of alloantibodies against HPA is essential step in the diagnosis of conditions such as fetal and neonatal alloimmune thrombocytopenia, post-transfusion purpura, and platelet transfusion refractoriness. The monoclonal antibody immobilization of platelet antigens (MAIPA) assay is the gold standard assay for the detection of anti-HPA antibodies. The need for fresh platelets from HPA-genotyped donors for MAIPA is a limitation. Alternative cell sources such as in vitro generated megakaryocytes/platelets or non-platelet cell lines are under investigation to overcome this limitation.

Aims: In the current study, we developed a monoclonal antibody immobilization of megakaryocyte antigens (MAIMA) assay using in vitro generated megakaryocytes for the determination of anti-HPA alloantibodies.

Methods: CD34+ cells were isolated from buffy coats of HPA-genotyped blood donors and differentiated for 28 days in vitro into megakaryocytes. The performance of MAIMA was assessed using WHO reference reagents (HPA-1a, HPA-3a and HPA-5b) and patients’ sera with well-characterized anti-HPA antibodies.

Results: Optical density (OD) values measured with the WHO anti-HPA-1a reference reagent were similar for megakaryocytes and platelets. On the other hand, OD values for the WHO anti-HPA-3a reference reagent were lower in MAIMA than in MAIPA. Anti-HPA-5b antibodies were not detectable in MAIMA. Anti-HPA-1a antibodies were successfully detected in MAIMA in all clinical samples (n=7). OD values in MAIPA and MAIMA showed high correlation (r=0.96, p< 0.001). MAIMA was reactive for samples with anti-HPA-3a (n=2), but OD values were 33-50% lower in MAIMA than in MAIPA. MAIMA was reactive in 2 of 3 samples with anti-HPA-3b antibodies. All patient samples with anti-HPA-5b antibodies were negative in MAIMA (n=4).

Conclusion(s): In conclusion, in vitro generated megakaryocytes showed potential to replace donor platelets, however, they might not be suitable for the detection of antibodies against certain HPA such as HPA-5b.

Learning Objectives:

Define the shortcomings of monoclonal antibody immobilization of platelet antigens assay.
Understand the importance of detecting alloimmune platelet antibodies in the management of fetal-neonatal alloimmune thrombocytopenia.
define the limitations of in vitro generated megakaryocytes in the detection of HPA alloantibodies.