Characterization of primary biliary cirrhosis (PBC) specific mitochondrial determinants by immunoblotting.

Antimitochondrial antibodies (AMA) are detected in up to 100% of patients with primary biliary cirrhosis (PBC); subtypes of AMA are disease specific. Sera from 21 patients with PBC and from 50 patients with various hepatic and non-hepatic diseases were tested for AMA by indirect immunofluorescence, radioimmunoassay for PBC-specific subtype anti M2 and characterized by western blot analysis and agarose-IEF-immunoblotting. Sera from patients with PBC reacted with up to 7 different mitochondrial polypeptides on western blots, mol. wt. 24,000-62,000 dalton. Sera from 50 patients with various hepatic and non-hepatic diseases did not react with these polypeptides. Sera with other AMA subtypes were included in this study (anti M1, anti M3, and anti M5). These mitochondrial polypeptides were associated with inner mitochondrial membranes (mitoplasts). Sonification led to a solubilization of several mitochondrial polypeptides (p 62, p 48, p 40, p 24). On agarose-IEF-immunoblotting sera from patients with PBC and 3 sera from patients with AMA positive cholestatic CAH but no other sera reacted with a protein band at pI 4.0; seven PBC sera reacted in addition with a protein band at pI 4.4. Western blot and agarose-IEF-immunoblotting are sensitive and specific tools to identify and characterize mitochondrial target antigens in PBC. Furthermore these techniques allow to study the clinical relevance of the heterogeneity of AMA in cholestatic liver disease.