Clinical chemistry and laboratory medicine · 2018
Reference standards for the detection of anti-mitochondrial and anti-rods/rings autoantibodies.
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Calise S John, Zheng Bing, Hasegawa Tomoko, Satoh Minoru, Isailovic Natasa, Ceribelli Angela, Andrade Luis E C, Boylan Katherine, Cavazzana Ilaria, Fritzler Marvin J, de la Torre Ignacio Garcia, Hiepe Falk, Kohl Kathryn, Selmi Carlo, Shoenfeld Yehuda, Tincani Angela, Chan Edward K L, IUIS Autoantibody Standardization Committee
Аннотация
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.
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.