Merlin Wilson

International recommendations for the assessment of autoantibodies to cellular antigens referred to as anti-nuclear antibodies

Anti-nuclear antibodies (ANA) are fundamental for the diagnosis of autoimmune diseases, and have been determined by indirect immunofluorescence assay (IIFA) for decades. As the demand for ANA testing increased, alternative techniques were developed challenging the classic IIFA. These alternative platforms differ in their antigen profiles, sensitivity and specificity, raising uncertainties regarding standardisation and interpretation of incongruent results. Therefore, an international group of experts has created recommendations for ANA testing by different methods. Two groups of experts participated in this initiative. The European autoimmunity standardization initiative representing 15 European countries and the International Union of Immunologic Societies/World Health Organization/Arthritis Foundation/Centers for Disease Control and Prevention autoantibody standardising committee. A three-step process followed by a Delphi exercise with closed voting was applied. Twenty-five recommendations for determining ANA (1–13), anti-double stranded DNA antibodies (14–18), specific antibodies (19–23) and validation of methods (24–25) were created. Significant differences between experts were observed regarding recommendations 24–25 (p<0.03). Here, we formulated recommendations for the assessment and interpretation of ANA and associated antibodies. Notably, the roles of IIFA as a reference method, and the importance of defining nuclear and cytoplasmic staining, were emphasised, while the need to incorporate alternative automated methods was acknowledged. Various approaches to overcome discrepancies between methods were suggested of which an improved bench-to-bedside communication is of the utmost importance. These recommendations are based on current knowledge and can enable harmonisation of local algorithms for testing and evaluation of ANA and related autoantibodies. Last but not least, new more appropriate terminologies have been suggested.

Immune reactants in cryoproteins. Relationship to complement activation.

Cryoproteins were isolated from the serum of 5 patients with essential cryoglobulinaemia 5 patients with rheumatoid arthritis, and 2 patients with Sjøgrens syndrome. These cryoprecipitates contained IgG, IgM, and IgA as well as complement proteins C1q, C4, C3, and factor B. The cryoprecipitates were analysed further for content of antibody and antigen, and were tested for their ability to activate complement. In the cryoprecipitates of 2 patients with Sjøgrens syndrome, nuclear antigen and antinuclear antibody characteristic of an immunological specificity found in Sjøgrens syndrome were shown. The cryoprecipitates of 6 other patients contained rheumatoid factor and antibody to a lymphocyte nuclear antigen. The solubilized cryoprecipitates were tested by in vitro assays for their ability to activate complement by the classical or alternative pathways. All 12 cryoprecipitates activated the classical pathway. 9 of the 12 cryoprecipitates also activated the alternative complement pathway under conditions which did not involve activation of C1 and C4. These studies show that a high percentage of cryoprecipitates consist at least in part of immune reactants. We discuss the relationship of these findings to pathogenetic mechanisms in disease.