PRODUCTION OF RABBIT POLYCLONAL ANTIBODY AGAINST APOBEC-1 BY GENETIC IMMUNIZATION

Circulating apolipoprotein B (apoB) exists in two forms; apoB-100 and apoB-48. ApoB-48 is a truncated form of apoB resulting from RNA editin g. The editing enzyme, called apobec-1, converts a cytidine (C) at nuc leotide 6666 in apoB-100 mRNA to a uridine (U) and changes a CAA codon to an in-frame stop codon, UAA. We have produced a specific rabbit po lyclonal antiserum against apobec-1 by genetic immunization. The cDNA of mouse apobec-1 was inserted down-stream and in-frame at the BamH I site in the last exon of human growth hormone cDNA driven by a cytomeg alovirus promoter. This plasmid was injected together with another pla smid expressing granulocyte macrophage colony-stimulating factor into the thigh muscles of a rabbit. The resulting antiserum demonstrated hi gh specificity on Western blots, and inhibited the apoB mRNA editing a ctivity of mouse liver extract in a dose-dependent manner. This report demonstrates that DNA immunization is a powerful technique that can b e readily applied to other sparse or difficult-to-purify proteins in l ipid metabolism.