Characterization of agglutinating monoclonal antibody against human red blood cells

Abstract

As a first step to clone cDNAs encoding variable domains of the heavy and the light chain of hemagglutinating antibody, mouse monoclonal antibodies against human red blood cells were generated and the N-termini of the heavy and the light chain were sequenced. Ficoll-paque purified red blood cells were used as a antigen to generate hybridoma and nine clones were selected as positive clones by a hemagglutination assay. One clone (# 12) was selected as a strong antibody producer and monoclonal antibody secreted from the clone was purified to homogeneity. By amino acid analysis, N-terminal sequence of the heavy chain was shown to be Glu-Val-Gln-Leu-Gln-Gln-Ser-Gly-Pro and that of the light chain was Asp-He-Val-Met-Thr-Gln-Ser-Pro. The heavy chain sequence was identified as antibody subgroup Ha and that of the light chain was subgroup I or subgroup HI. Taken together, these results provided useful information to design oligonucleotides for cloning of antibody cDNA which is initial phase work toward developing immunodiagnostic using human whole blood.