Identification of specific antinuclear antibodies in dogs using a line immunoassay and enzyme-linked immunosorbent assay

Abstract

Circulating antinuclear antibodies (ANA) are commonly present in the systemic autoimmune disease Systemic Lupus Erythematosus (SLE) and in other systemic rheumatic diseases, in humans as well as in dogs. The indirect immunofluorescence (IIF)-ANA test is the standard method for detecting ANA. Further testing for specific ANA with immunoblot techniques or ELISAs is routinely performed in humans to aid in the diagnosis and monitoring of disease. Several specific ANA identified in humans have been identified also in suspected canine SLE but, in contrast to humans, investigation of autoantibodies in canine SLE is mainly restricted to the IIF-ANA test. Our aim was to identify both known and novel specific ANA in dogs and to investigate if different IIF-ANA patterns are associated with different specific ANA in dogs. Sera from 240 dogs with suspicion of autoimmune disease (210 IIF-ANA positive (ANApos) and 30 IIF-ANA negative (ANAneg)) as well as sera from 27 healthy controls were included. The samples were analysed with a line immunoassay, LIA (Euroline ANA Profile 5, Euroimmun, Lübeck, Germany) and four different ELISAs (Euroimmun). The ANApos dogs were divided in two groups depending on the type of IIF-ANA pattern. Of the 210 ANApos samples 68 were classified as ANA homogenous (ANAH) and 141 as ANA speckled (ANAS), one sample was not possible to classify. Dogs in the ANAH group had, compared to the other groups, most frequently high levels of anti-double stranded deoxyribonucleic acid (dsDNA) and anti-nucleosome ANA. Anti-dsDNA antibodies were confirmed in some dogs with the Crithidia luciliae indirect immunofluorescence test (CLIFT). The frequency of ANAH dogs with values above those observed in the healthy group was significantly higher compared to ANAS dogs for anti-dsDNA, anti-nucleosome, and anti-histone reactivity. Dogs in the ANAS group had, compared to the other groups, most frequently high levels of anti-ribonucleoproteins (RNP) and/or anti-Smith (Sm) antibodies. Reactivity against Sjögren's syndrome related antigens (SS)-A (including the Ro-60 and Ro-52 subcomponents), SS-B, histidyl tRNA synthetase (Jo-1), topoisomerase I antigen (Scl-70), polymyositis-scleroderma antigen (PM-Scl) and proliferating cell nuclear antigen (PCNA) was also noted in individual dogs. In conclusion, by using a commercial LIA and different ELISAs originally developed for detection of human ANA, we identified several specific ANA in serum samples from dogs sampled for IIF-ANA testing. Further, we found that the types of IIF-ANA pattern were associated with reactivity against some particular nuclear antigens.