Detection of antimitochondrial autoantibodies in immunofluorescent AMA-negative patients with primary biliary cirrhosis using recombinant autoantigens

Abstract

Antimitochondrial antibodies (AMA) are the serologic hallmark of primary biliary cirrhosis (PBC). However, depending on the clinical laboratory, from 5% to 17% of PBC patients are consistently AMA-negative, using native mitochondrial antigens and a variety of conventional assays including immunofluorescence (IMF) and enzyme-linked immunosorbent assay (ELISA). The major immunoreactive mitochondrial autoantigens are the E2 members of the 2-oxo-acid dehydrogenase complex family, including pyruvate dehydrogenase complex-E2 (PDC-E2), branched chain 2-oxo acid dehydrogenase complex-E2 (BCOADC-E2), and oxo-glutarate dehydrogenase complex-E2 (OGDC-E2); cDNAs of these proteins have now been cloned, sequenced, and their B-cell epitopes defined. In the present study, we cloned cDNAs encoding these proteins from human, not bovine, sources, and expressed the recombinant proteins in a newly developed ELISA that employs a unique Escherichia coli buffer, and compared the data with previous assays using both AMA-positive and -negative patients. Using this new assay and our criteria for positive as an optical density (OD) greater than 10 SD above the mean of control sera, the AMA-positive rate of 191 PBC sera was 94% (

Antimitochondrial antibodies in acute liver failure: Implications for primary biliary cirrhosis

“…In studies of AMA reactivity in large numbers of sera, we have demonstrated that when recombinant antigens are used for immunodiagnosis, they are strikingly specific for PBC; we have also shown that such specificity and sensitivity is dramatically increased with the use of ELISA and/or immunoblot assays compared with immunofluorescence. 21,24,25 In our initial analysis and our observations that AMAs were indeed found in ALF, we decided to replicate the data in a second site-hence the duplicate analysis at both UC Davis and Teikyo University (both sites had identical results). Recent work has suggested that posttranslational modification of the mitochondrial 2-OADC lipoyl moiety is a possible mechanism in the initiation of AMAs in PBC.…”

Risk factors and comorbidities in primary biliary cirrhosis: A controlled interview-based study of 1032 patients

“…Patients were eligible for inclusion if the diagnosis of PBC had been made between 1996 and 2004 and was based on pre-determined criteria, including a detectable AMA and at least one of the following: a cholestatic pattern of serum biochemical tests (serum alkaline phosphatase Ͼ2 times normal upper values) for at least 6 months and/or compatible liver histology. 12 In suspected cases of PBC without detectable AMA (up to 10% of patients in the routine clinical laboratory 13 ), criteria were modified and included all the following: ANA or anti-smooth muscle autoantibodies, a cholestatic pattern of biochemical tests (elevated serum alkaline phosphatase) for at least 6 months, and a compatible liver histology. To assess accuracy, referring physicians re-evaluated anonymized clinical information in 100 randomly selected enrolled patients.…”

Liver-targeted and peripheral blood alterations of regulatory T cells in primary biliary cirrhosis

“…AMAs were detected through the use of recombinant mitochondrial antigens. 24 All patients, including AMA-negative subjects, fulfilled the diagnostic criteria of PBC based on internationally accepted standards and were all undergoing ursodeoxycholic acid treatment. 2 Patients who did not have fibrosis at the time of liver biopsy are considered as stages I or II PBC according to Ludwig et al, 25 while those showing signs of fibrosis or cirrhosis at histology were considered as stages III or IV (Table 1).…”