Quantification of platelet-bound immunoglobulins of different class and subclass using radiolabeled monoclonal antibodies: assay conditions and clinical application
G. J. Tijhuis
Central Laboratory of the Netherlands Red Cross Blood Transfusion Service and Laboratory for Experimental and Clinical Immunology, University of Amsterdam, Amsterdam, and Department of Haematology, Academic Medical Centre, University of Amsterdam, Amsterdam, The Netherlands
Search for more papers by this authorR. J. L. Klaassen
Central Laboratory of the Netherlands Red Cross Blood Transfusion Service and Laboratory for Experimental and Clinical Immunology, University of Amsterdam, Amsterdam, and Department of Haematology, Academic Medical Centre, University of Amsterdam, Amsterdam, The Netherlands
Search for more papers by this authorP. W. Modderman
Central Laboratory of the Netherlands Red Cross Blood Transfusion Service and Laboratory for Experimental and Clinical Immunology, University of Amsterdam, Amsterdam, and Department of Haematology, Academic Medical Centre, University of Amsterdam, Amsterdam, The Netherlands
Search for more papers by this authorW. H. Ouwehand
Central Laboratory of the Netherlands Red Cross Blood Transfusion Service and Laboratory for Experimental and Clinical Immunology, University of Amsterdam, Amsterdam, and Department of Haematology, Academic Medical Centre, University of Amsterdam, Amsterdam, The Netherlands
Search for more papers by this authorCorresponding Author
A. E. G. Kr. vondem Borne
Central Laboratory of the Netherlands Red Cross Blood Transfusion Service and Laboratory for Experimental and Clinical Immunology, University of Amsterdam, Amsterdam, and Department of Haematology, Academic Medical Centre, University of Amsterdam, Amsterdam, The Netherlands
Dr A. E. G. Kr. von dem Borne, c/o Publication Secretariat, Central Laboratory of the Netherlands Red Cross Blood Transfusion Service, P.O. Box 9406, 1006 AK Amsterdam, The Netherlands.Search for more papers by this authorG. J. Tijhuis
Central Laboratory of the Netherlands Red Cross Blood Transfusion Service and Laboratory for Experimental and Clinical Immunology, University of Amsterdam, Amsterdam, and Department of Haematology, Academic Medical Centre, University of Amsterdam, Amsterdam, The Netherlands
Search for more papers by this authorR. J. L. Klaassen
Central Laboratory of the Netherlands Red Cross Blood Transfusion Service and Laboratory for Experimental and Clinical Immunology, University of Amsterdam, Amsterdam, and Department of Haematology, Academic Medical Centre, University of Amsterdam, Amsterdam, The Netherlands
Search for more papers by this authorP. W. Modderman
Central Laboratory of the Netherlands Red Cross Blood Transfusion Service and Laboratory for Experimental and Clinical Immunology, University of Amsterdam, Amsterdam, and Department of Haematology, Academic Medical Centre, University of Amsterdam, Amsterdam, The Netherlands
Search for more papers by this authorW. H. Ouwehand
Central Laboratory of the Netherlands Red Cross Blood Transfusion Service and Laboratory for Experimental and Clinical Immunology, University of Amsterdam, Amsterdam, and Department of Haematology, Academic Medical Centre, University of Amsterdam, Amsterdam, The Netherlands
Search for more papers by this authorCorresponding Author
A. E. G. Kr. vondem Borne
Central Laboratory of the Netherlands Red Cross Blood Transfusion Service and Laboratory for Experimental and Clinical Immunology, University of Amsterdam, Amsterdam, and Department of Haematology, Academic Medical Centre, University of Amsterdam, Amsterdam, The Netherlands
Dr A. E. G. Kr. von dem Borne, c/o Publication Secretariat, Central Laboratory of the Netherlands Red Cross Blood Transfusion Service, P.O. Box 9406, 1006 AK Amsterdam, The Netherlands.Search for more papers by this authorAbstract
Summary. Quantification of platelet-bound immunoglobulins (PBIg) with radiolabelled murine monoclonal antibodies (mAbs) has been described only for IgG so far. Here we describe some modifications of this mAb radioimmunoassay (MARIA) and show that by using a panel of radiolabelled specific mAbs it is possible to quantify not only PBIgG but also PBIgG subclasses and PBIgM. Analysis by gel filtration showed that all anti-IgG and anti-IgG-subclass mAbs bound to their respective antigens in a ratio of about 1:1. However, the binding ratio for the anti-IgM Mab could not be established. There was a good correlation between the antibody-density per platelet as determined with the anti-IgG mAb and determined as the sum of the IgG molecules of different subclass per platelet (r= 0·90). Platelet fragments did not interfere in the assay. 89 normal healthy controls had 140 IgG molecules per platelet and bound 269 anti-IgM molecules per platelet (geometric means).
In a study on the detection of PBIg in 147 thrombocytopenic patients, it appeared that the MARIA had a sensitivity of 61% and a specificity of 45% for the diagnosis of idiopathic thrombocytopenic purpura (ITP). Both in ITP and in secondary thrombocytopenia (STP), PBIgG1 and PBIgG3 were found more frequently (60% and 61%, respectively) than PBIgG2and PBIgG4 (13% and 9%, respectively). There was no relation between the amount of total PBIgG or PBIgM and the platelet count in either ITP or STP. Also, if IgG antibodies of only one subclass were found, there was no relation between the severity of the thrombocytopenia and the amount of PBIgG.
By applying the MARIA, it is possible to quantify PBIgG, all four PBIgG-subclasses and PBIgM in ITP and STP in a reliable way.
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