Cloning and functional analysis of anti-double strand DNA IgG autoantibodies using the phage-display method.

To examine the relationship between pathophysiological effects and molecular features of anti-double-stranded (ds)DNA autoantibodies (Abs), we isolated anti-dsDNA Ab fragments by using the phage-display method. Fd ç and light-chain DNA were PCR amplified from peripheral blood lymphocytes of a patient with systemic lupus erythematosus (SLE), complicated with lupus nephritis. They were then inserted into a phagemid vector, pComb3-H. We generated eight Fab fragments that specifically bound to solid-phase DNA. The Fab clones stained positively using Crithidia luciliae, indicating that they were anti-dsDNA antibodies. Nucleotide sequences of VH of Fab clones were very similar and appeared to be derived from VH26 germline gene. Differences in the activities of anti-dsDNA Ab of the Fab clones may be ascribed to the diversity of VL. Fab fragments with anti-dsDNA Ab activities exhibited a positive charge on isoelectric focusing, consistent with pathogenic features. Our results indicate that anti-dsDNA Ab Fab fragments obtained by the phage-display method in the present study may possess molecular and functional characteristics of pathogenic anti-dsDNA autoAbs in SLE.