This is because the currently used massive parallel DNA sequencing approaches do not easily identify all the structural variations in the RCA gene cluster. We will describe here how to use the MLPA assays and two computational tools to analyze NGS data, NextGENe and ONCOCNV, to detect CNVs and gene rearrangements in the RCA gene cluster.C3 nephritic Factor (C3NeF) is autoantibody that binds neoepitopes of the C3 convertase C3bBb, resulting in a stabilization of the enzyme. First functional characterizations of C3NeF were performed by hemolytic assays using preactivated sheep erythrocytes (bearing C3b). Sheep erythrocytes are beforehand sensitized with an anti-sheep red blood cell stroma antibody produced in rabbit (hemolysin). Sensitized sheep erythrocytes will initiate cascade complement activation via the classic pathway, followed by alternative pathway amplification loop, resulting in C3b covalent binding to cell surface. Sheep erythrocytes bearing C3b permit the alternative pathway exploration, in particular decay of alternative pathway C3 convertase.Antibodies to autoantigens are implicated in a large number of diseases. Such autoantibodies may cause pathological activation of complement, an ancient humoral recognition and effector system of innate immunity; in addition, complement components or regulators may be target of autoantibodies and cause abnormal complement activation or function. Autoantibodies to complement proteins are in particular involved in kidney diseases. Those binding to complement convertase enzymes can cause enhanced stability of convertases and their increased resistance to regulation, thus promoting complement turnover. Here, we describe an ELISA method to detect factor B autoantibodies that bind to and stabilize the alternative complement pathway C3 convertase enzyme, C3bBb.Autoantibodies against complement proteins are involved in the pathological process of many diseases, including lupus nephritis, C3 glomerulopathies, and atypical hemolytic uremic syndrome. This method describes the detection of autoantibodies targeting the central complement component C3 by ELISA. These autoantibodies (IgG) are detected in up to 30% of the patients with lupus nephritis and more rarely in cases with C3 glomerulopathies. These autoantibodies recognize the active fragment C3b and have overt functional consequences. They enhance the formation of the C3 convertase and prevent the inactivation of C3b by Factor H and complement receptor 1. Moreover, they enhance the deposition of complement activation fragments on activator surfaces, such as apoptotic cells. The data currently available on the relations of anti-C3 autoantibodies with clinical, laboratory, and histological markers for activity of lupus nephritis, as well as the relations of anti-C3 with classical immunological markers for activity of autoimmune process in patients with lupus nephritis, such as hypocomplementemia and high levels of anti-dsDNA, could identify these autoantibodies as a potential marker for evaluation the activity of lupus nephritis. These autoantibodies correlate with the disease severity and can be used to identify patients with lupus nephritis who were prone to flare. Therefore, the detection of such autoantibodies could guide the clinicians to evaluate and predict the severity and to manage the therapy of lupus nephritis.Ficolins are recognition proteins of the lectin pathway of the complement system and also play an important role in innate immunity and in the maintenance of tissue homeostasis. They deserve special attention in the context of autoimmunity since they are involved in the uptake of dying cells. Because the monitoring of systemic lupus erythematosus (SLE) patients is particularly difficult, it is crucial to find new relevant serum biomarkers. The ability to detect autoantibodies in the patients' sera provides a diagnostic and prognostic advantage. We describe in this chapter quantitative enzyme linked immunosorbent assays (ELISA) to detect the presence of autoantibodies targeting ficolin-2 and ficolin-3 in human sera.  https://www.selleckchem.com/products/blu-451.html  Recombinant ficolins produced in a mammalian expression system are used as coating antigens. The described in-house ELISAs provide a valuable tool to efficiently quantify anti-ficolin autoantibodies in the sera of SLE patients.Enzyme-linked immunosorbent assay (ELISA) is a quantitative analytical method used to measure the concentration of molecules in biological fluids through antigen-antibody reactions. Here we describe the measurement of anti-C1-inhibitor autoantibodies by an indirect ELISA. In this method patients' sera are incubated in a microplate coated with plasma derived C1-inhibitor.Autoantibodies against complement C1q (anti-C1q) are an excellent marker for active nephritis in SLE patients. Here, we describe a typical protocol for the quantification of anti-C1q using immobilized C1q (important for the presentation of relevant cryptic epitopes) and a high salt buffer for the incubation steps (to prevent immune-complex binding to intact C1q). More recently, a linear epitope on the C1q A chain, that is targeted by anti-C1q, has been described (A08). The assay using this peptide seems to be more specific and more sensitive for the detection of active nephritis in SLE patients than the conventional anti-C1q assay, but further studies are required to establish the role of anti-A08 of C1q in the clinical routine.The three pathways of the complement system converge toward the cleavage of the central complement component C3 into its activated fragments, with C3b being able to bind covalently to the activating surface. The endothelial cells are among the major targets for complement attack in pathological conditions, as the atypical hemolytic uremic syndrome. Therefore, study of complement C3 deposition on endothelial cells by flow cytometry is a sensitive test to measure complement activation. This test can be used as a research or clinical tool to test complement activation induced by patients' sera or to test the functional consequences of newly discovered complement mutations as well as different triggers of endothelial cells injury.