Reactivity of sera from patients with
primary biliary cirrhosis (PBC) with a 60 kDa component of nuclear pore complexes (NPCs), purified by affinity chromatography on
wheat-germ agglutinin (
WGA)-Sepharose, was previously detected. Recently, clinical significance of the anti-NPC
antibodies in PBC became evident. In the light of recent reports, indicating the correlation of the anti-NPC
antibodies with severity and progression of the disease, the characterization of the reactive
antigens is becoming essential in the clinical management of patients with PBC. Since accurate
autoantibody detection represents one of the fundamental requirements for a reliable testing, we have generated a human recombinant p62
protein and validated an immunoprecipitation assay for the detection of anti-p62. We also demonstrated that the generated human recombinant p62
nucleoporin was modified by
N-acetylglucosamine residues. More than 50% of tested PBC sera precipitated (35)S-radioactively labeled p62 recombinant
nucleoporin and 40% recognized this recombinant
antigen by immunoblotting. We compared the reactivity of PBC sera with rat and human
nucleoporin. The incidence of anti-p62
nucleoporin positive PBC sera increased by 15% when human recombinant
antigen was used. The titer of
autoantibodies in p62-positive PBC samples strongly varied. Preadsorption of the PBC sera with p62
recombinant protein completely abolished their reactivity with the
antigen. In conclusion, this study unequivocally proves that
autoantibodies reacting with the 60 kDa component of NPCs target p62
nucleoporin and, more importantly, provide a better
antigen source for future evaluations of the clinical role of anti-p62 in PBC.