Identification of a human heavy chain antibody fragment directed against human platelet alloantigen 1a by phage display library.
The human platelet alloantigen HPA-1a (PlA1) is responsible for most cases of post-transfusion purpura and neonatal alloimmune thrombocytopenia in the Caucasian population. HPA-1a and HPA-1b are two allelic forms of the platelet membrane glycoprotein IIIa (GPIIIa) gene that differ by a single amino acid. In this report, we describe the development of a recombinant heavy chain antibody fragment capable of distinguishing between the homozygous forms of HPA-1a and HPA-1b. This antibody fragment was isolated from the lymphocytes of an immunized individual through the use of a phage display library system. The recombinant antibody fragment reacted with human platelet lysates from HPA-1a homozygous donors, the HPA-1a form of recombinant N-terminal GPIIIa and intact HPA-1a platelets, but did not react with platelet lysate from HPA-1b homozygous donors, reduced HPA-1a form of platelet GPIIIa or other platelet glycoproteins. This HPA-1a specific human antibody fragment works well in common laboratory assays such as ELISA and flow cytometry, which can assist in identifying HPA-1b homozygous individuals who are known to have a higher risk for developing neonatal alloimmmune thrombocytopenia and post-transfusion purpura. Thus, selection of recombinant antibody fragment using phage display offers a promising alternative to hybridoma technology for the production of human antibodies against human alloantigens and holds potential as a technique in therapeutic applications.
Okamoto
N
N
Biomedical Research Center, Olympus Optical Co., Ltd., Hachiojishi, Tokyo, Japan.
Kennedy
S D
SD
Barron-Casella
E A
EA
Casella
J F
JF
Inoko
H
H
Kickler
T S
TS
D000918:Antibody Specificity; D016824:Antigens, Human Platelet; D017104:Bacteriophage M13 / Q000235:genetics; D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D015723:Gene Library; D006801:Humans; D007128:Immunoglobulin Fragments / Q000235:genetics*; D007143:Immunoglobulin Heavy Chains / Q000235:genetics*; D039661:Integrin beta3; D007518:Isoantibodies / Q000235:genetics; D007519:Isoantigens; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology
D016824:Antigens, Human Platelet; D017931:DNA Primers; C061248:ITGB3 protein, human; D007128:Immunoglobulin Fragments; D007143:Immunoglobulin Heavy Chains; D039661:Integrin beta3; D007518:Isoantibodies; D007519:Isoantigens; D011994:Recombinant Proteins; C083451:human platelet antigen 1b