title: Sensitive detection of platelet-specific antibodies with a modified MAIPA using biotinylated antibodies and streptavidin-coated beads.
abstract: We have developed a modified monoclonal antibody immobilization of platelet antigens assay (MAIPA) with enhanced sensitivity in detecting antibodies against human platelet antigens (HPA), using biotinylated monoclonal antibodies, streptavidin-coated beads and detection by flow cytometry. The beads-MAIPA gave superior signal-to-noise resolution (>10-fold higher) for detection of anti-HPA-1a and anti-HPA-5b compared with the in-house standard MAIPA. Also, efficient and reproducible detection of anti-HPA-15 (CD109) was shown. The enhanced sensitivity was confirmed using WHO International Reference Reagents for anti-HPA-1a, anti-HPA-3a and anti-HPA-5b, which allowed comparison of detection endpoints with other laboratories. Finally, the beads-MAIPA was validated for quantification of anti-HPA-1a. The lower limit of quantification was 0.4IU/mL for beads-MAIPA, compared to 1IU/mL previously reported for standard MAIPA. Based on improved performance against all HPA-antibodies tested, the beads-MAIPA has replaced the standard MAIPA in our laboratory in diagnostics of conditions due to HPA-immunization, such as fetal and neonatal alloimmune thrombocytopenia (FNAIT).
authors.affiliation: Dept. of Clinical Immunology and Transfusion Medicine, Karolinska University Hospital, Stockholm, Sweden; Dept. of Laboratory Medicine, Karolinska Institute, Stockholm, Sweden.
authors.forename: Anette
authors.initials: A
authors.lastname: Mörtberg
authors.affiliation: Dept. of Clinical Immunology and Transfusion Medicine, Karolinska University Hospital, Stockholm, Sweden; Dept. of Medicine Huddinge, Center for Hematology and Regenerative Medicine (HERM), Stockholm, Sweden.
authors.forename: Stephan
authors.initials: S
authors.lastname: Meinke
authors.affiliation: Dept. of Clinical Immunology and Transfusion Medicine, Karolinska University Hospital, Stockholm, Sweden.
authors.forename: Petra
authors.initials: P
authors.lastname: Berg
authors.affiliation: Dept. of Laboratory Medicine, University Hospital of North Norway, Tromsø, Norway.
authors.forename: Mette Kjær
authors.initials: MK
authors.lastname: Killie
authors.affiliation: Dept. of Laboratory Medicine, University Hospital of North Norway, Tromsø, Norway.
authors.forename: Jens
authors.initials: J
authors.lastname: Kjeldsen-Kragh
authors.affiliation: Prophylix Pharma AS, Tromsø Science Park, Tromsø, Norway.
authors.forename: Kerstin
authors.initials: K
authors.lastname: Järås
authors.affiliation: Dept. of Laboratory Medicine, Karolinska Institute, Stockholm, Sweden.
authors.forename: Erle
authors.initials: E
authors.lastname: Refsum
authors.affiliation: Dept. of Clinical Immunology and Transfusion Medicine, Karolinska University Hospital, Stockholm, Sweden; Dept. of Medicine Huddinge, Center for Hematology and Regenerative Medicine (HERM), Stockholm, Sweden.
authors.forename: Petter
authors.initials: P
authors.lastname: Höglund
authors.affiliation: Dept. of Clinical Immunology and Transfusion Medicine, Karolinska University Hospital, Stockholm, Sweden; Dept. of Laboratory Medicine, Karolinska Institute, Stockholm, Sweden. Electronic address: agneta.wikman@karolinska.se.
authors.forename: Agneta
authors.initials: A
authors.lastname: Wikman
mesh_terms: D000911:Antibodies, Monoclonal / Q000737:chemistry; D016824:Antigens, Human Platelet / Q000097:blood* / Q000145:classification; D019921:Biotinylation; D001792:Blood Platelets / Q000737:chemistry; D005434:Flow Cytometry; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D016014:Linear Models; D008863:Microspheres; D012680:Sensitivity and Specificity; D019809:Streptavidin / Q000737:chemistry; D054098:Thrombocytopenia, Neonatal Alloimmune / Q000175:diagnosis
chemical_list: D000911:Antibodies, Monoclonal; D016824:Antigens, Human Platelet; D019809:Streptavidin
keywords: Alloimmunisation; Anti-HPA-1a quantification; FNAIT; Human platelet antigen; MAIPA; Platelet antibody