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ANA testing by line immunoassay – the present scenario

It has been more than 50 years when anti nuclear antibodies were first discovered and found to be associated with connective tissue diseases. And since then many tests have been discovered and used for their detection and confirmation. For many decades , ANA IFA has been considered and continued to be considered as the Gold standard method for ANA testing. However to increase sensitivity and specificity of the test further approaches were discovered.

In case of a suspected CTD, the clinicians check for a positive ANA result in conjunction with clinical and diagnostic findings. However, since different ANA are associated with one or other CTD a systematic approach has to be followed while performing these tests. Therefore initially screening is carried out usually by IF-ANA/ELISA and if positive, more specific tests are performed based on clinical findings and IF-ANA staining patterns.

Line blot Immunoassay was discovered in 1980 and is qualitative and more specific test which offers antibody reactivity to antigens that are applied as distinct separate lines on an immunochromatographic membrane.

Many manufactures today offer an array of different ANA line blot immunoassay however the antigen specificities taken into testing panel differ. The most common antigens taken for ANAtesting by Line blot assay shall be able to confirm the most common CTDs like Systemic Lupus Erythmatosus, Sjogren’s syndrome, Scleroderma, CREST, Myositis (PM/DM), Systemic sclerosis and PBC.

Offering different panels for ANA testing with different antigens could be very confusing for test interpretation and thus it is very helpful when a line blot immunoassay offers major ANAs on a single strip.

Myositis autoantibodies are traditionally categorized in two groups, based on their diagnostic accuracy: myositis-specific antibodies (MSA) and myositis-associated antibodies (MAA), the latter mostly occurring in myositis-overlap syndromes. Besides the other traditional MSA like Jo-1, the new markers for Myositis are still being missed in these tests. Anti-Mi-2 antibodies are newly conceived immune targets that have been recently identified, mostly in patients with severe forms of dermatomyositis or necrotizing myopathy. The prevalence rate of these antibodies has been found to be 18%.

Among the MAA, anti-PM/Scl and anti-Ku characterize an overlap polydermatomyositis/systemic sclerosis ( PM/SSc) syndrome with severe interstitial lung involvement. The prevalence rate of these antibodies are 9 and 5% respectively.

These autoantibodies are useful for predicting clinical manifestations, treatment outcomes, and vital prognoses.

Apart from this, the test should be able to offer the most pure form of antigens like dsDNA for SLE detection and avoiding any false positive result in dsDNA testing. Many a times the impurities on the band while coating antigens cause lot of background colour after testing. The test thus shall offer clear background for increased specific results.

The test interpretation is again a concern with ANA line blot immunoassay. The interpretation software should be easy to use and disallows any manipulation in results for accurate diagnosis.

To know more on how to set ANA blot in your lab setting, click here.

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