Abstract
Antinuclear antibody (ANA) is useful in the diagnosis of connective tissue disorder (CTD). Association of specific autoantibodies with the immunofluorescence pattern of ANA in CTD, noted in western literature has been considered as reference in all over the world. However, in Bangladesh no such research work or data correlating the autoantibodies and their ANA patterns is found. Objective of the study was to identify an association between immunofluorescence patterns of antinuclear antibody on HEp-2 cell and more specific antinuclear reactivities (e.g. anti-dsDNA and anti-extractable nuclear antigen) in the serum samples of CTD patients. Serum samples of 152 CTD patients (Systemic lupus erythematosus, Rhumatoid arthritis, Sjogren's syndrome, Systemic sclerosis, Polymyositis, Mixed connective tissue disease) were diagnosed clinically, attending at Bangabandhu Sheikh Mujib Medical University (BSMMU) during the study period of January, 2010 to December, 2010. Samples were subjected for ANA testing by Indirect Immunofluorescence (IIF) on HEp-2 cell (ALPHADIA) in dilution of 1:40, anti-dsDNA by ELISA and anti- extractable nuclear antigen (anti-ENA) by Dot Immunoblot. Dot blot strips were tested for anti-Sm, anti-RNP, anti-SSA/Ro, anti-SSB/La, anti-Scl-70 and anti-Jo-1. Out of 152 patients 110 (72.3%) cases were ANA positive by IIF on HEp-2 cell. ANA positive sera exhibited four fluorescence patterns such as speckled (50.8%), peripheral (21.6%) , homogenous (18.1%) and nucleolar pattern (9%). Peripheral pattern and homogenous pattern was predominantly associated with anti-dsDNA (p < 0.05). Speckled pattern was significantly associated with anti-ENA (p < 0.05).The most commonly identified antinuclear autoreactivity was directed towards anti-RNP (25.7%) then anti-Scl-70 (20%), anti-SSA (14.2%) and anti-SSB (5.7%). Multiple anti-ENA reactivities were identified in 34.28% cases. Peripheral and homogenous pattern is strongly associated with anti-dsDNA and speckled pattern may predict anti-ENA (specially ribonucleoprotiens). As a definite correlation between the ANA patterns and the group of antibodies was detected by dot immunoblot, one could predict presence of certain specific auto antibodies for a particular ANA pattern identified. This may restrict on the cost of laboratory investigations in a developing country like Bangladesh. Thus, ANA-IIF method may reduce the expense of detailed immunological work-up with minimal loss in diagnostic accuracy.
Highlights
An autoimmune response is a common manifestation of the connective tissue diseases
Four patterns of nuclear fluorescence were noted: the homogenous pattern, in which the entire nucleus fluoresced (Fig.1); the peripheral pattern, in which the fluorescence is located along the rim of nucleus (Fig.2); the speckled pattern in which the fluorescence is localized as discrete spots in the nucleus (Fig.3); and the nucleolar pattern in which fine and discrete speckles in the nucleolus within a dark border (Fig.4)
In these fluorescence positive samples, speckled pattern was the most common pattern seen in 50.9% cases, followed by peripheral pattern 21.8%, homogenous pattern 18.2% and nucleolar pattern 9.1%
Summary
Antinuclear antibody (ANA) test and tests for specific autoantibodies to nuclear antigen play an important role in the diagnostic evaluation, prognostic assessment and monitoring of patients with autoimmune connective tissue disorder (CTD).[1] The indirect immunofluorescence (IIF) performed on cultured human epithelial (HEp-2) cell is currently considered to be the gold standard for the detection of antinuclear antibodies (ANA). For detection of immune profile, additional techniques such as western immunoblotting, enzyme linked immunosorbent assay (ELISA) or line immunoassay are required.[2,3,4] Since a characteristic profile of ANA is associated with most CTD, identification of the fine specificity may provide valuable clues to the diagnosis These are usually inferred using data obtained from western studies. No such study has been done in the Bangladeshi population
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