These results demonstrate that this C-terminal PRYSPRY domain of Ro52 contains useful epitopes for the diagnosis of SjS, but that these antibodies are of lower titer than the N-terminus and are missed by ELISAs which use recombinant proteins. Open in a separate window Figure 3 LIPS detection of autoantibodies against Ro60, Ro52 and a C-terminal fragment of Ro52 in 25 normal volunteers and 57 primary SjS patients. autoantibody attack of thyroid, neuronal and gastric parietal cells, respectively. These heterogeneous autoantibody responses detected by LIPS in SjS will likely be useful for diagnosis and for evaluating extraglandular manifestations. Keywords: Autoantibodies, Autoantigen, Diagnosis Sj?grens syndrome, SSA, SSB Introduction Sj?grens syndrome (SjS) is a common autoimmune disorder associated with epithelial inflammation and exocrine glands dysfunction [1]. The 2-Hydroxybenzyl alcohol etiology of SjS is usually unknown and genetic and environmental factors are both thought to play a role in triggering the autoimmune response. The presence of multiple autoantibodies and oligoclonal and polyclonal B cell activation is usually characteristic of SjS. The spectrum of SjS extends from an organ-specific autoimmune disease with sicca symptoms of dry eyes and mouth, to systemic disease with diverse extraglandular manifestations involving other organs such as the thyroid, lungs, gastrointestinal and the peripheral nervous system. The current American-European consensus classification for primary SjS is based 2-Hydroxybenzyl alcohol on six criteria [2]. Indicators of ocular and oral dryness, evidence of inflammation from minor salivary gland biopsy, and the presence of autoantibodies towards extractable nuclear SSA and SSB antigens are included in the criteria. These antibodies are not specific to SjS, however, as they are detectable in SLE (systemic lupus erythematosus), myositis and other autoimmune diseases [3]. SSA is composed of structurally unrelated Ro52 and Ro60, while SSB, also known as La, is a single 48 kDa protein. In the 2002 classification standards for SjS diagnosis, positive SSA and SSB autoantibody titers was the only mandatory criteria for primary SjS if the salivary gland biopsy was unfavorable [2]. Current ELISA-based immunoassays detect a rate of approximately 70% SSA and 40% SSB immunoreactivity in SjS [1]. However, few autoantibody assessments exist to evaluate the diverse extraglandular manifestations that are commonly found in SjS. Most ELISA-based immunoassays for evaluating autoantibodies, including those for SSA and SSB, employ either native proteins or protein 2-Hydroxybenzyl alcohol complexes, such as those purified from calf thymus [4], or ones produced recombinantly in bacteria or baculovirus. Immunoassays that utilize proteins from these sources may contain 2-Hydroxybenzyl alcohol contaminating proteins or macromolecular aggregates that limit the accuracy of these assessments due to high backgrounds and/or false signals. We recently developed a highly sensitive immunoprecipitation technology called Luciferase Immunoprecipitation System (LIPS) that utilizes mammalian cell-produced, recombinant fusion protein antigens for efficiently evaluating antibody responses [5C11]. In a limited number of side-by-side Rabbit Polyclonal to NCoR1 comparisons, LIPS showed improved diagnostic performance compared to existing immunoassays for detecting antibodies to infectious brokers [10, 11] and provided new tools to monitor drug treatment [11] and sub-stratify disease says [9]. In addition, LIPS is usually highly useful for profiling autoimmunity [7,8] and in one study showed several advantages over a highly sensitive radioactive transcription/translation assay for detecting anti-IA2 autoantibodies associated with type I diabetes [8]. In the present study, LIPS was evaluated for its diagnostic performance in detecting autoantibodies to SSA and SSB and for screening additional autoantibody responses in primary SjS and control volunteers. Material and methods Patients and methods The sera were randomly selected from a cohort of patients with primary SjS participating in a longitudinal natural history study. These sera included 57 well-characterized patients diagnosed with primary SjS and 25 healthy volunteers evaluated under Institutional Review Board-approved protocols at the SjS clinic of the National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD. An additional 15 serial samples from the primary SjS patients were tested for further validation of the LIPS tests but were not used for calculations of sensitivity and specificity. As part of a clinical.
