Reference standards for the detection of anti-mitochondrial and anti-rods/rings autoantibodies

Clin Chem Lab Med. 2018 Sep 25;56(10):1789-1798. doi: 10.1515/cclm-2017-1152.


Background:: Anti-mitochondrial antibodies (AMA) are found in >90% of primary biliary cholangitis patients. Anti-rods/rings antibodies (anti-RR) are most commonly associated with interferon-α and ribavirin treatment in hepatitis C patients. Clinical laboratories routinely screen for AMA and anti-RR using indirect immunofluorescence on HEp-2 cells (HEp-2-IFA). Therefore, we sought to establish reference materials for use in AMA and anti-RR testing.

Methods:: AMA-positive and anti-RR-positive human plasma samples (AMA-REF and RR-REF), identified as potential reference materials based on preliminary data, were further validated by multiple laboratories using HEp-2-IFA, immunoprecipitation (IP), western blotting, IP-western, line immunoassay (LIA), addressable laser bead immunoassay (ALBIA), and ELISA.

Results:: AMA-REF showed a strong positive cytoplasmic reticular/AMA staining pattern by HEp-2-IFA to ≥1:1280 dilution, and positive signal on rodent kidney/stomach/liver tissue. AMA-REF reacted with E2/E3, E3BP, E1α, and E1β subunits of the pyruvate dehydrogenase complex by IP and western blotting, and was positive for AMA antigens by LIA, ALBIA, and ELISA. RR-REF showed a strong positive rods and rings staining pattern by HEp-2-IFA to ≥1:1280 dilution. RR-REF reacted with inosine monophosphate dehydrogenase by IP, IP-western, and ALBIA. RR-REF also produced a nuclear homogenous staining pattern by HEp-2-IFA, immunoprecipitated proteins associated with anti-U1RNP antibody, and reacted weakly with histones, nucleosomes, Sm, and nRNP/Sm by LIA.

Conclusions:: AMA-REF and RR-REF are useful reference materials for academic or commercial clinical laboratories to calibrate and establish internal reference standards for immunodiagnostic assays. AMA-REF and RR-REF are now available for free distribution to qualified laboratories through Plasma Services Group.

Keywords: anti-mitochondrial antibody; anti-rods/rings antibody; autoantibody; autoimmunity; hepatitis C; primary biliary cholangitis.

MeSH terms

  • Aged
  • Antiviral Agents / adverse effects
  • Antiviral Agents / therapeutic use
  • Autoantibodies / blood*
  • Autoantibodies / immunology
  • Autoimmune Diseases / diagnosis
  • Female
  • Fluorescent Antibody Technique, Indirect / standards
  • Hepatitis C / drug therapy
  • Humans
  • Immunoassay / methods*
  • Immunoassay / standards
  • Immunoprecipitation
  • Interferon-alpha / adverse effects
  • Interferon-alpha / therapeutic use
  • Laboratories
  • Middle Aged
  • Mitochondria / immunology*
  • Reference Standards
  • Ribavirin / adverse effects
  • Ribavirin / therapeutic use


  • Antiviral Agents
  • Autoantibodies
  • Interferon-alpha
  • Ribavirin