Working Algorithms and Detection Methods of Autoantibodies in Autoimmune Liver Disease: A Nationwide Study

Abstract

Autoantibody detection is the cornerstone of autoimmune liver diseases (AILD) diagnosis. Standardisation of working algorithms among autoimmunity laboratories, as well as being aware of the sensitivity and specificity of various commercial techniques in daily practice, are still necessary. The aim of this nationwide study is to report the results of the 2020 Autoimmunity Workshop organised by the Autoimmunity Group of the Spanish Society of Immunology and to provide useful information to clinicians and laboratory specialists to improve the management of autoantibody detection in AILD diagnoses. Serum samples from 17 patients with liver diseases were provided by the organisers of the 2020 Autoimmunity Workshop and were subsequently analysed by the 40 participating laboratories. Each laboratory used different techniques for the detection of autoantibodies in each patients’ serum sample, according to their working algorithm. Thus, almost 680 total complete patient reports were obtained, and the number of results from different autoantibody detection techniques was >3000. Up to eight different working algorithms were employed, including indirect immunofluorescence assays (IFA) and antigen-specific techniques (AgST). The IFA of HEp-2 cells was more sensitive than IFA of rat triple tissue for the study of anti-nuclear autoantibodies (ANA) associated with AILD. The IFA of a human neutrophil study for the analysis of anti-neutrophil cytoplasmic autoantibodies was not carried out systemically in all patients, or by all laboratories. AgSTs were the most sensitive methods for the detection of anti-smooth muscle/F-actin, soluble liver antigen, liver cytosol-1, M2-mitochondrial autoantibodies, and ANA associated with primary biliary cholangitis. The main differences in AMA detection were due to patients with autoantibodies against the non-dominant epitope of pyruvate dehydrogenase complex. Given that they are complementary, IFA and AgST should be performed in parallel. If there is high suspicion of AILD, AgST should always be performed.

Detection of M2-Type Anti-Mitochondrial Autoantibodies against Specific Subunits in the Diagnosis of Primary Biliary Cholangitis in Patients with Discordant Results

“…Our group previously published a study in which sera of 17 patients with liver diseases were analyzed by 40 laboratories, each with its diagnostic algorithm. For some patients, a low percentage of laboratories reported AMA-M2 pattern by IFA; however, this percentage was clearly superior when samples were analyzed by antigen-specific techniques using the three E2 subunits together or separately [6].…”

“…Primary biliary cholangitis (PBC) is a chronic autoimmune liver disorder characterized by inflammation of intrahepatic biliary ducts, which leads to fibrosis and consequently may produce cirrhosis and liver failure [1][2][3][4][5]. Its prevalence is low and it mostly affects middle-aged women [1,6]. The etiology is unclear, but many data suggest that a genetic predisposition along with environmental risk factors such as infections or smoking may be a crucial responsible [1,4].…”

“…AMA-M2 target components of the 2-oxo-acid dehydrogenase complex: pyruvate dehydrogenase complex (PDC), 2-oxoglutarate dehydrogenase complex (OGDC) and branched-chain 2-oxoacid dehydrogenase complex (BCOADC). Specifically, autoantibodies mainly recognize the E2 subunits of these complexes: PDC-E2 (80-90% of cases), BCOADC-E2 (50-80% of cases) and OGDC-E2 (20-60% of cases), and to a lesser extent, the E1 and E3 subunits [2,5,6].…”

“…The accurate detection of AMA-M2 is relevant as it is considered the hallmark of the disease. Its recognition could avoid a biopsy and lead to an earlier diagnosis, which is essential to start an adequate treatment [5,6]. Traditionally, IFA has been the most accepted assay for its detection; however, several antigen-specific methods such as ELISA or Dotblot (DB) have been developed.…”

“…Traditionally, IFA has been the most accepted assay for its detection; however, several antigen-specific methods such as ELISA or Dotblot (DB) have been developed. These assays use a fusion protein combining the three E2 subunits, a mixture of recombinant E2 subunits or the three E2 recombinant subunits isolated, among others [5,6,8].…”