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A novel method for simultaneous analysis of specific platelet antibodies: SASPA
Author(s) -
Nguyen Xuan Duc,
Dugrillon Alex,
Beck Christian,
Kerowgan Mohammad,
Klüter Harald
Publication year - 2004
Publication title -
british journal of haematology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.907
H-Index - 186
eISSN - 1365-2141
pISSN - 0007-1048
DOI - 10.1111/j.1365-2141.2004.05233.x
Subject(s) - antibody , platelet , monoclonal antibody , microbiology and biotechnology , antigen , platelet membrane glycoprotein , flow cytometry , primary and secondary antibodies , immunology , monoclonal , chemistry , biology
Summary Glycoprotein (GP)‐specific platelet antibodies can cause allo‐immune and auto‐immune thrombocytopenia. The specific detection of relevant antibodies is a prerequisite for diagnosis and treatment. Here, we describe an improved method based on simultaneous detection of various platelet‐specific immunoglobulin G (IgG) and IgM antibodies. Bead populations with distinct fluorescence intensities, coated with monoclonal antibodies specific for mouse heavy chain isotypes, were used for the simultaneous immobilization of platelet‐GP [IIb/IIIa, Ib/IX, human leucocyte antigen (HLA) class I, or Ia/IIa, CD32, GPIV or CD109, Ib/IX, HLA class I]. In order to detect human IgG and IgM antibodies simultaneously, phycoerythrin‐ and fluorescein isotiocyanate‐conjugated goat anti‐human IgG and IgM were added. On this basis, the abundance of six distinct antibodies (three anti‐GP, each with subclasses IgG and IgM) were simultaneously analysed without cross‐reaction by flow cytometry. For evaluation, sera and platelets from 169 patients with platelet‐binding and/or platelet‐associated antibodies were investigated. The monoclonal antibody‐specific immobilization of platelet antigen (MAIPA) assay was performed in parallel as reference test. The simultaneous analysis of platelet‐specific antibodies (SASPA) assay was able to detect all platelet‐specific IgG and IgM that were also recognized by MAIPA with a comparable sensitivity. SASPA proved to be a rapid and reliable assay that required less platelets than other methods. This method has the potential to pave the way for new investigations of platelet‐specific antibodies.