Detection of anti-DNA topoisomerase I antibody by an enzyme-linked immunosorbent assay using overlapping recombinant polypeptides

Clin Immunol Immunopathol. 1995 Sep;76(3 Pt 1):266-78. doi: 10.1006/clin.1995.1125.


Five recombinant fusion proteins with overlapping amino acid sequences encompassing the entire DNA topoisomerase I (topo I) molecule were generated, purified, and used as antigens for an enzyme-linked immunosorbent assay (ELISA). IgG, IgA, IgM, and "total" (total of IgG, IgA, and IgM isotypes) anti-topo I antibodies were measured using a mixture of these five fusion proteins in 73 systemic sclerosis (SSc) sera positive for anti-topo I antibody by double immunodiffusion (DID) and 184 control sera negative for anti-topo I antibody by DID. Fragment-specific anti-topo I antibodies were also measured by ELISA using each topo I recombinant protein as antigen. IgG, IgA, IgM, and total anti-topo I antibodies were detected in 67 (92%), 56 (77%), 16 (22%), and 70 (96%) of 73 SSc sera positive for anti-topo I antibody by DID, respectively. The specificity of the total anti-topo I ELISA was 99% when compared with DID. The total anti-topo I ELISA levels were strongly correlated with DID titers (r = 0.907, P < 0.0001). Three sera which recognized a conformational epitope on native topo I or had predominantly IgM anti-topo I antibody showed a false-negative result with the total anti-topo I ELISA. Three SSc sera negative for anti-topo I antibody by DID were positive by the total anti-topo I ELISA, and two were confirmed to recognize the N-terminus of topo I. IgG and IgA antibody levels to the N-terminal and central portion of topo I were correlated with each other, but those to the C-terminus were not. These findings indicate that the ELISA using recombinant fusion proteins is a highly sensitive and specific alternative to conventional DID for the detection of anti-topo I antibody.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adult
  • Aged
  • Antibodies, Antinuclear / classification
  • Antibodies, Antinuclear / immunology*
  • DNA Topoisomerases, Type I / immunology*
  • DNA Topoisomerases, Type I / isolation & purification
  • Enzyme-Linked Immunosorbent Assay / methods
  • Female
  • HeLa Cells
  • Humans
  • Immunodiffusion
  • Immunoglobulin A / analysis
  • Immunoglobulin G / analysis
  • Immunoglobulin Isotypes / blood
  • Immunoglobulin M / analysis
  • Male
  • Middle Aged
  • Peptides
  • Recombinant Fusion Proteins / isolation & purification


  • Antibodies, Antinuclear
  • Immunoglobulin A
  • Immunoglobulin G
  • Immunoglobulin Isotypes
  • Immunoglobulin M
  • Peptides
  • Recombinant Fusion Proteins
  • DNA Topoisomerases, Type I