coli strain carrying a cloned cDNA encoding the allergen component of interest

coli strain carrying a cloned cDNA encoding the allergen component of interest. Testing with and without competitive CCD inhibition was performed within the same run and day on ImmunoCAP in order to measure the rate of false-positives due to CCD-sIgE binding to the cellulose solid phase. carbohydrate determinant (CCD) structures found in plant and insect glycoproteins are commonly recognized by IgE antibodies as epitopes that can lead to extensive cross-reactivity and obscure in vitro diagnostic (IVD) serology results. With the introduction of component resolved diagnosis (CRD), recombinant non-glycosylated components have been utilized to mitigate the risk of CCD-specific IgE (sIgE) detection. However, a recent study has shown that CCD-sIgE may bind directly to the cellulose solid phase matrix used in certain diagnostic assays, eliminating the advantage of CRD over traditional extract-based testing. The aim of this study is to further investigate the prevalence of CCD-sIgE interference on a commonly-used sIgE automated platform which employs a cellulose-based matrix to immobilize CCD-free recombinant components. Methods Sera from patients sensitized to peanut, silver birch, and/or timothy grass were analyzed for CCD-sIgE reactivity on ImmunoCAP/Phadia and NOVEOS autoanalyzers against the MUXF3 carbohydrate component. Positive CCD-sIgE sera were further analyzed against non-glycosylated recombinant components bound to the ImmunoCAP solid phase in the absence and presence of a soluble CCD inhibitor. For comparison, sera were then analyzed on NOVEOS, a non-cellulose based automated sIgE assay. Results Sera from 35% of the sensitized population tested in this study were positive (0.35 kU/L) for CCD-sIgE. Of those positives, 17% resulted in CCD-sIgE-positive (false positive) results on ImmunoCAP using non-glycosylated allergosorbents that were negative on NOVEOS. Sera producing false-positive results on ImmunoCAP had varying levels of CCD-sIgE from 0.67 kU/L to 36.52 kU/L. Rat monoclonal to CD4.The 4AM15 monoclonal reacts with the mouse CD4 molecule, a 55 kDa cell surface receptor. It is a member of the lg superfamily,primarily expressed on most thymocytes, a subset of T cells, and weakly on macrophages and dendritic cells. It acts as a coreceptor with the TCR during T cell activation and thymic differentiation by binding MHC classII and associating with the protein tyrosine kinase, lck The incidence of CCD interference was predominantly delimited to low-positive IgE results (0.35 kUA/LC 3.00 kUA/L). Conclusion Falsely elevated diagnostic allergen-sIgE results can commonly occur due to the presence of CCD-sIgE using assays that employ a carbohydrate matrix-based allergosorbent. Even the use of non-glycosylated recombinant allergenic components coupled to cellulose matrices do not reduce their risk of detection. The risk of CCD interference that compromises quantitative IgE results can be mitigated by the addition of a soluble CCD inhibitor to positive CCD-sIgE containing sera or by alternatively using a non-cellulose based sIgE assay, such as the NOVEOS assay. Introduction Glycoproteins found in plants and insects display structural homology across taxonomically diverse allergenic sources due to the presence of complex asparagine-linked oligosaccharides known as N-glycans.[1C3] More specifically, it is the presence of a core 1,3-linked fucose or a 1,2-linked xylose that represent common post-translational modifications of glycoproteins in these species and are the key elements of the widespread carbohydrate pan-epitope structure.[1,4C7] N-glycans with this epitope are known as cross-reactive carbohydrate determinants (CCDs) which contain core modifications that differ from those found in human glycoproteins. Thus, these can be viewed by the human immune system as foreign and, in some individuals, may elicit the production of IgE antibodies.[1] IgE antibodies reactive with CCD epitopes are believed to have limited or no clinical significance partly due to their low avidity and marginal biological activity.[8,9] When detected by diagnostic (IVD) assays, CCD reactive-IgE antibodies neither predicts the development of Pim1/AKK1-IN-1 clinical symptoms upon allergen exposure nor does it associate with disease severity.[10C12] CCD reactivity, however, can impact the diagnostic accuracy of the quantitative measurement of IgE antibodies in a patients serum analysis. Approximately 30% of the allergic population sera contain CCD-sIgE.[13,14] Component resolved diagnosis (CRD), using recombinant allergens with no apparent glycosylation, has therefore been recommended to reduce the risk of obtaining inaccurate results.[15,16] CRDs ability to discriminate between various aspects of clinical disease results in an improved diagnostic specificity and sensitivity. This leads to more effective therapeutic strategies and accurate predictions of allergic disease severity.[1,17C19] Currently, the most widely used single complexity allergen-specific IgE assay utilizing CRD is the ImmunoCAP (Thermo Scientific, ImmunoDiagnostics, Uppsala, Sweden) with over 100 components available for testing. However, a recent study has shown that the ImmunoCAP polymerized cellulose matrix used to bind allergenic proteins contains CCD epitopes that are recognizable by IgE antibodies.[20] This means that Pim1/AKK1-IN-1 the CCD-sIgE of a patient tested against an advertised CCD-free recombinant protein, such as rAra h 8, may Pim1/AKK1-IN-1 recognize N-glycans present on the cellulose matrix; which would result in an increased rate of false-positive results, and at a minimum, reduce confidence in the accuracy of results generated on these cellulose-based assays. With many clinicians unaware of this issue, the interpretation of recombinant CRD results may lead to an incorrect diagnosis if the patient sera contains levels of CCD-sIgE. The aim of this study was to further investigate the prevalence of CCD interference on the ImmunoCAP allergen-specific IgE assay when utilizing CCD-free recombinant proteins and to offer suitable alternatives for specific IgE testing when the need to mitigate the detection of CCD-sIgE is required. Materials and methods Study population In total, 204 serum samples were selected for this study based on a positive ( 0.35 kUA/L) sIgE result to peanut, silver birch, and/or.