The purpose of the present study is to evaluate the centromeric pattern on human laryngeal tumour (HEp-2) cells by indirect immunofluorescent (IIF) test and to compare their reactivities with a newly developed recombinant centromere protein B enzyme linked immunosorbent assay (CENP-B ELISA) test using sera of antinuclear antibody (ANA)-reactive primary biliary cirrhosis (PBC) patients. Antimitochondrial antibody (AMA) subtypes (PDC-E2, BCOADC-E2, OGDC, protein X, and PDC-E1 alpha) by Western blot were also investigated to see whether they have any effect on the expression of CENP-B reactivities. A centromeric pattern (anticentromere antibody [ACA]) was detected in 11 of 25 (44%) PBC patients whereas CENP-B reactivity was found in 15 (60%) of them. There were some differences in IIF patterns and CENP-B reactivities. One PBC serum with indistinguishable ANA pattern reacted with CENP-B. Eight of 15 (53%) CENP-B reactive patients had other autoimmune-like disorders. Of 181 healthy sera, none was reactive for ACA either by IIF or by ELISA test. There was a correlation between ACA IIF and CENP-B ELISA titres (r = 0.824, P < 0.001). However, no correlation was observed between either CENP-B or AMA reactivities and/or between either autoantibodies or laboratory and histologic indices of PBC. These findings suggest that recombinant CENP-B ELISA appears to be more sensitive in identifying ACA than IIF, underlying its potential value as a screening test for the diagnosis of PBC complicated with other autoimmune-like disorders. The presence of multiple autoantibodies in PBC sera may reflect heterogeneous antigens recognition, and requires further study to identify target antigens at cellular and molecular levels.