An antibody against single-stranded DNA useful for detecting apoptotic cells : production and applications for development, differentiation and carcinogenesis

点击翻译按钮获取中文摘要

In this research project we developed an improved method for the production of the antibody and investigated into the antigenic determinants of the antibody so that it could be of practical use for detecting apoptotic cells. Rabbits, hyperimmunized with complexes of alkaline-denatured calf thymus DNA and methylated bovine serum albumin, produced an IgG antibody to single-stranded DNA.Analysis by sandwich ELISA using various naturally occurring nucleic acids revealed that the antibody was specific to single-stranded DNA.Furthermore, using synthetic polymers in the assay, it was found that the antibody could recognize single-stranded DNA with various base sequences. Gel electrophoresis retardation assays, with synthetic oligodeoxynucleotides with differing lengths of single-stranded DNA,indicated that a hexadeoxynucleotide constituted the minimum size of the antigenic determinants, and suggested that the antibody probably consists of several antibodies which recognize hexadeoxynucleotides with various base sequences. Western blot analysis demonstrated that the antibody can recognize both a DNA ladder and oligonucleosomes prepared from rat liver nuclei with endogenous endonuclease. We also demonstrated that this antibody is a useful tool for detecting apoptotic cells in development, differentiation and carcinogenesis.