autoantibodies in serum or plasma bind to the antigen-coated surface. The wells are then washed to remove unbound components. In the second incubation, the enzyme-labeled polyclonal antibody to an immunoglobulin (conjugate) binds with surface-bound autoantibodies. After additional washing, specific autoantibodies are traced with substrate during incubation. Addition of a stop solution terminates the reaction, which results in a coloured end product. The amount of conjugate bound is measured in absorbance units (Wild, 2013, p. 9).
These assays can be quantified through high throughput specialized machinery such as the IMMAGE 800 Immunochemistry System and/or the AU Chemistry Analyzers (various models) manufactured by Beckman Coulter…show more content… 3).
The RF immunosorbent assay can be used in conjunction with another biomarker to develop a whole other assay. One of the two examples presented in Egerer’s journal article (2009) uses RF and mutated citrullinated vimentin (MCV) antibodies to develop the Rheuma-Check test and the second example shows development of the CCPoint assay by using antibodies to CCP (p. 162). The two serological tests are considered point-of-care tests (POCT) because they aid in the determination for early detection of RA (Egerer, 2009, p. 162). In order for these analytes to be used and manipulated for assay purposes, one must understand the biological and immunological properties. Furthermore, other potential POCTs can regard filaggrin as a citrullinated antigen, which can be expressed in keratin-producing epithelial cells for detection in assays by cross-reacting with anti-keratin antibodies (AKA) (Egerer, 2009, p. 160). The “posttranslational modification” process of citrullination includes alterations of the protein charge, which distorts the structure and antigenic properties (Egerer, 2009, p. 160). Also, understanding the sensitivity and specificity of the binding process also makes for better assay-POCT development and adaptation. The sensitivity for RA follows that the diagnostic properties of filaggrin and citrullinated fibrin in rheumatoid arthritis are comparable to the CCP antigen (Egerer, 2009, p. 160). Moreover,