PB1110 - Buffy-coat-derived megakaryocytes for monoclonal antibody immobilization of platelet antigens (MAIMA) assay

Tuesday, July 12, 2022

6:30 PM – 7:30 PM

Location: Exhibition

Presentation Format: In-Person

Presenting Author(s)

Günalp Uzun, MD

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

Co-Author(s)

JL

Josip Lucic

Doctor
Department of Transfusion Medicine, Cellular Therapeutics and Hemostaseology, Clinic for Anesthesiology LMU University Hospital
Munich, 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
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
SN

Stefanie Nowak-Harnau

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

Irene Marini

Post Doc
Institute for Clinical and Experimental Tranfusion Medicine
Tuebingen, 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

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

Background: Serologic detection and characterization of anti-human platelet antigen (HPA) antibodies are essential for proper diagnosis and treatment in fetal and neonatal immune thrombocytopenia. The monoclonal antibody immobilization of platelet antigens (MAIPA) assay is the standard assay for the detection of anti-HPA antibodies. For this assay, fresh platelets from HPA-genotyped donors are required, which is a limitation for laboratories with no direct access to blood donors.

Aims: In the current study, the use of buffy coat-derived, in-vitro cultured megakaryocytes (MKs) instead of donor-derived platelets in MAIPA for the determination of anti-HPA antibodies was investigated.

Methods: We isolated CD34+ cells from buffy coats of blood donors using magnetic beads. CD34+ cells were differentiated for 28 days in vitro into megakaryocytes. Megakaryocyte cell lines were characterized with flow cytometry using CD41, CD61, CD42a and CD42d expression and DNA content. The performance of MAIPA with megakaryocytes was assessed using WHO reference sera und patients’ sera with well-characterized anti-HPA antibodies.

Results: Testing WHO standard sera in MAIPA with megakaryocytes (MAIMA) revealed similar reaction compared to standard MAIPA using platelets (OD HPA 1a:0.51±0.13 vs. 0.52±0.20, p=0.99 and OD for HPA5b (0.15±0.09 vs. 0.25±0.07, p=0.40, Figure 1). However, OD amplitudes were significantly lower in MAIMA than in MAIPA with donor platelets for HPA 3a (0.19±0.03 vs. 0.53±0.07, p=0.02, Figure 1). OD amplitudes were similar with megakaryocytes and platelets in MAIPA using patients’ sera with anti-HPA-1a and anti-HPA 5b antibodies (Figure 2).

Conclusion(s): Buffy-coat-derived megakaryocytes have the potential to replace donor platelets in MAIPA in the future. Further studies are needed for standardization and validation of this technique in routine laboratories.