Anti-SS-A/Ro
Enzyme-linked immunosorbent assay method for the semi-quantitative determination of specific IgG autoantibodies to SS-A/Ro in human serum.
The detection of Anti-Nuclear Antibodies (ANA’s) has long been an important tool in the diagnosis of systemic rheumatic diseases. The antigens used in their detection are purified by the saline extraction of human or animal nuclei, this has led to them being termed Extractable Nuclear Antigens (ENA’s). The most commonly measured ENA specifications are anti-SS-A/Ro, anti-SS-B/La, anti-Sm, anti-Sm/RNP, anti-Jo-1 and anti-Scl-70.
The intracellular antigen SS-A/Ro is a target for autoimmune response for many patients with Systemic Lupus Erythematosus (SLE), Sjogren’s Syndrome (SS) and other connective tissue diseases. Studies have shown that anti-SS-A/Ro antibodies occur in 60-70% of patients with primary Sjogren’s Syndrome and 30-50% of patients with SLE, being particulary associated with photosensitive dermatitis. The autoantibodies are also often found in the majority of patients with subacute cutaneous lupus erythematosus or antinuclear antibody negative SLE. The SS-A/Ro antigen has been shown to be a trypsin sensitive ribonucleoprotein which under Western Blot produces 2 bands at 52KD and 60KD.
The AutostatII assay for detection of autoantibodies is a solid phase immunosorbent assay (ELISA) in which the analyte is indicated by a colour reaction of an enzyme and substrate. The AutostatII wells are coated with purified antigen (1).
On adding diluted serum to the wells the antibodies (2) present bind to the antigen. After incubating at room temperature and washing away unbound material, horseradish peroxidase conjugated anti-IgG monoclonal antibody (3) is added, which binds to the immobilised antibodies.
Following further incubation and washing, tetra-methyl benzidine substrate (TMB) (4) is added to each well. The presence of the antigen-antibody-conjugate complex turns the substrate to a dark blue colour. Addition of the stop solution turns the colour to yellow.
The colour intensity is proportional to the amount of autoantibodies present in the original serum sample.