Identification of novel clusters of co-expressing cytokines in a diagnostic cytokine multiplex test.

Introduction: Cytokines are mediators of the immune system that are essential for the maintenance, development and resolution of immune responses. Beneficial immune responses depend on complex, interdependent networks of signaling and regulatory events in which individual cytokines influence the production and release of others. Since disruptions in these signaling networks are associated with a wide spectrum of diseases, cytokines have gained considerable interest as diagnostic, prognostic and precision therapy-relevant biomarkers.

Microbead-based technologies in diagnostic autoantibody detection.

Background: There is a rapid proliferation of new technologies to identify a spectrum of autoantibodies in medical conditions that range from organ-specific autoimmune diseases to systemic rheumatic diseases. Although many laboratories have adopted high-throughput diagnostic platforms such as enzyme linked immunoassays (ELISA), other technologies such as microbead-based assays are emerging as an alternative diagnostic platform.

Objective: To understand the performance and importance of bead based immunoassays in clinical diagnostics and therapeutics.

Identification of GW182 and its novel isoform TNGW1 as translational repressors in Ago2-mediated silencing.

RNA interference is triggered by small interfering RNA and microRNA, and is a potent mechanism in post-transcriptional regulation for gene expression. GW182 (also known as TNRC6A), an 182-kDa protein encoded by TNRC6A, is important for this process, although details of its function remain unclear. Here, we report a novel 210-kDa isoform of human GW182, provisionally named trinucleotide GW1 (TNGW1) because it contains trinucleotide repeats in its mRNA sequence.

The emergence of multiplexed technologies as diagnostic platforms in systemic autoimmune diseases.

In the last decade, there has been a rapid proliferation of new technologies that are capable of identifying an increasing spectrum of autoantibodies and other biomarkers in autoimmune diseases. These newer diagnostic technologies include line immunoassays, addressable laser bead immunoassays, microarrays in microfluidics platforms and nanobarcode particles. Multiplexed bead assays are a particularly robust platform because they are adaptable to the detection of a variety of disease specific biomarkers, such as autoantibodies, cytokines, adipokines, drugs, oligonucleotides and single nucleotide polymorphisms, Although many laboratories have adopted a variety of these diagnostic platforms to improve turn around times and meet budget constraints, there is an urgent need to ensure that the rapid adoption of new technologies is attended by an appropriate balance of assay sensitivity and specificity.

Analysis of human sera that are polyreactive in an addressable laser bead immunoassay.

Background: Following the introduction of addressable laser bead immunoassays (ALBIA) into our clinical laboratory, it was noted that certain sera would exhibit reactivity to numerous antigens in the array. To further understand the nature of this reactivity, we analyzed the reactivity of sequential sera that were identified over a 1 year period.

Methods: Sera that demonstrated reactivity to 6 or more of the 8 antigens in an ALBIA kit (QuantaPlex 8: chromatin, Sm, RNP, Scl-70, ribosomal P protein, SS-A/Ro, SS-B/La, Jo-1) were tested for autoantibodies by indirect immunofluorescence (IIF) on HEp-2 cell substrates, for IgG, IgM and IgA rheumatoid factor, chromatin and ribosomal P protein by ELISA and by LINE immunoassay (LIA) and immunoblotting (IB).

Identification of the B-cell epitopes of the early endosome antigen 1 (EEA1).

Early endosome antigen 1 (EEA1) is a target autoantigen in patients diagnosed with neurological and other autoimmune conditions. Eighteen of 65 sera (28%) that displayed a vesicular cytoplasmic staining pattern also immunoprecipitated the recombinant EEA1. These 18 sera were selected for further clinical, serological and epitope mapping studies.

A panel of monoclonal antibodies to cytoplasmic GW bodies and the mRNA binding protein GW182.

GW182 is a mRNA binding protein characterized by 60 repeats of glycine (G):tryptophan (W) motifs and is localized in cytoplasmic structures referred to as GW bodies (GWBs). Current evidence suggests that this unique protein plays a role in mRNA processing. To enable a more detailed study of GW182 and GWBs in cells and tissues, including their role in mRNA processing, we developed four monoclonal antibodies (MAbs) that bind the human recombinant GW182 protein.

The use and abuse of commercial kits used to detect autoantibodies.

The detection of autoantibodies in human sera is an important approach to the diagnosis and management of patients with autoimmune conditions. To meet market demands, manufacturers have developed a wide variety of easy to use and cost-effective diagnostic kits that are designed to detect a variety of human serum autoantibodies. A number of studies over the past two decades have suggested that there are limitations and concerns in the use and clinical application of test results derived from commercial kits.