Establishment of a cell line panel for the detection of antibodies against human platelet antigen 4b

Abstract

Antibodies against human platelet antigens (HPAs) play important roles in thrombocytopenia. In Japan, HPA-4b antibody is frequently responsible for HPA-related neonatal alloimmune thrombocytopenia. A highly sensitive assay using platelets has been developed for the detection of antibodies against HPAs. However, it is difficult to obtain the platelets expressing specific HPAs required for the assay. Therefore, an alternative method not requiring platelets would be helpful to detect antibodies against HPAs. Glycoprotein IIIa (GPIIIa) cDNA encoding HPA-4b was individually co-transduced with that of wild-type GPIIb in K562 cells, which is a non-adherent cell line, using a retroviral vector. The expression of transgene products in cultured cells were observed for over 6 months. Next, to evaluating the sensitivity and specificity of this cell line panel, we performed monoclonal antibody-specific immobilization of platelet antigens (MAIPA) assay with a serum previously identified by another method. All HPA-4b antibodies in serum samples were positive, and all serum samples, including normal serum and serum containing HLA antibodies were negative. No difference was observed in the specificity and sensitivity between our method and conventional MAIPA using platelets. The present results indicate that this established cell line panel permits highly sensitive detection of specific antibodies against HPA-4b.

Severe neonatal thrombocytopenia due to anti‐HPA‐4b alloantibodies: Third report described in a Caucasian mother

“…Luminex technology with Pak Lx assay identified an anti‐HPA‐4b alloantibody associated to class I HLA antibodies (MATCH IT! software interpretation) since we have no cell lines expressing HPA‐4b in the laboratory 9 …”

Establishment of a cell panel for detecting antibodies against human platelet antigen 2b located on CD42

“…Its specificity was determined using an anti-PLT-MPHA-Screen kit (Beckman Coulter, Tokyo, Japan). The MAIPA assay was performed according to the method described by Kiefel (1992), with minor modifications (Hayashi et al, 2011b), using a monoclonal anti-CD42b antibody, AK2. The results are expressed as optical densities at 450 nm.…”