Increasing Efficiency and Quality by Consolidation of Clinical Chemistry and Immunochemistry Systems with MODULAR ANALYTICS SWA

MODULAR ANALYTICS Serum Work Area (in USA Integrated MODULAR ANALYTICS, MODULAR ANALYTICS is a trademark of a member of the Roche Group) represents a further approach to automation in the laboratory medicine. This instrument combines previously introduced modular systems for the clinical chemistry and immunochemistry laboratory and allows customised combinations for various laboratory workloads. Functionality, practicability, and workflow behaviour of MODULAR ANALYTICS Serum Work Area were evaluated in an international multicenter study at six laboratories. Across all experiments, 236000 results from 32400 samples were generated using 93 methods. Simulated routine testing which included provocation incidents and anomalous situations demonstrated good performance and full functionality. Heterogeneous immunoassays, performed on the E-module with the electrochemiluminescence technology, showed reproducibility at the same level of the general chemistry tests, which was well within the clinical demands. Sample carryover cannot occur due to intelligent sample processing. Workflow experiments for the various module combinations, with menus of about 50 assays, yielded mean sample processing times of <38 minutes for combined clinical chemistry and immunochemistry requests; <50 minutes including automatically repeated samples. MODULAR ANALYTICS Serum Work Area offered simplified workflow by combining various laboratory segments. It increased efficiency while maintaining or even improving quality of laboratory processes

Effect of Heating and Glycation on the Allergenicity of 2S Albumins (Ara h 2/6) from Peanut

Although no effect of processing on T-cell reactivity was observed, heat induced denaturation reduced the IgE reactivity and subsequent functionality of Ara h 2/6. Conversely, Ara h 2 and 6 purified from roasted peanut retained the structure and IgE reactivity/functionality of the native protein which may explain the allergenic potency of this protein. Through detailed molecular study and allergenicity assessment approaches, this work then gives new insights into the effect of thermal processing on structure/allergenicity of peanut proteins

The Relevance of Nutrition for Pediatric Allergy and Immunity

The development of the immune system in early life is essential to shape an immune system [...

Immunological Characterization of Dutch Sesame Seed-Allergic Patients

Background: Sesame seed is an allergen of growing importance worldwide. However, knowledge of the clinically relevant sesame allergen and its cross-reactivity with homologous allergens is limited. The aim of this study was the immunological characterization of Dutch sesame seed-allergic patients and evaluation of cross-reactivity between sesame seed, tree nut and pollen allergens using different sources of allergen extracts. Methods: Six patients with a medical history of sesame seed allergy were included, i.e. 5 with an anaphylactic reaction and 1 with an oral allergy syndrome (OAS). The immunological background of the sesame seed and tree nut IgE sensitization was characterized with Western blotting and a basophil activation test (BAT). The major sesame allergen was identified by nanoLC-MS/MS. Cross-reactivity was measured using an immuno-inhibition assay with the Phadia ImmunoCAP system. Results: Oleosin was identified as the major allergen for the 5 patients with an anaphylactic reaction to sesame seed, but no cross-reactivity between sesame and tree nut proteins was observed. For the patient with OAS, IgE specific to oleosin was not detected but cross-reactivity between sesame seed and tree nut proteins was observed. The BAT and ImmunoCAP inhibition test added value to the clinical and immunological characterization of sesame seed-sensitized patients, distinguishing relevant and non-relevant sensitizations. Conclusions: Our immunological approach enabled us to fully characterize the sensitization pattern of 6 sesame seed-allergic patients. The different protein composition of commercially available allergen extracts influences the outcomes of the immunological assays and thus also the diagnosis to a large extent.</p

The Basophil Activation Test reduces the need for a food challenge test in children suspected of IgE-mediated cow's milk allergy

Background: The gold standard for the diagnosis of cow's milk allergy is the Double-Blind Placebo-Controlled Food Challenge (DBPCFC) test. However, disadvantages of the DBPCFC are the potential risk of anaphylactic reactions, the time-consuming procedure and high costs. Objective: The aim of this study was to determine the reliability of the Basophil Activation Test (BAT) both for the initial diagnosis of cow's milk allergy in children and for the determination of tolerance in children with cow's milk allergy. Methods: Ninety-seven BATs and cow's milk-specific IgE (sIgE) tests were performed in 86 infants/young children, suspected of (persistent) cow's milk allergy, who were qualified for an in-hospital DBPCFC. The BAT was performed with cow's milk extract and the purified major allergens casein, α-lactalbumin, β-lactoglubulin. Basophil activation was determined by CD63 upregulation measured by flow cytometry. The BAT results were compared to the DBPCFC outcomes. Results: Based on unequivocal DBPCFC and BAT result combinations (80%), the BAT had a sensitivity and specificity of 100% (CI: 86%-100% and 68%-100%, respectively) in IgE-sensitized children (41% of the tested children). All non-IgE-sensitized children (59%) had a negative DBPCFC and BAT, except for five patients. These latter showed delayed and relatively mild symptoms in the DBPCFC with a negative BAT, supporting a non-IgE-mediated allergy in these children. Conclusions and Clinical Relevance: The BAT seems reliable and cost-effective to diagnose patients with an IgE-mediated cow's milk allergy. In IgE-sensitized patients, a BAT might replace a DBPCFC. For non-IgE-sensitized patients presenting with mild symptoms, we propose to consider a (double-blind) extended (time) challenge test at home.</p

Receptor Mediated Effects of Advanced Glycation End Products (AGEs) on Innate and Adaptative Immunity : Relevance for Food Allergy

As of late, evidence has been emerging that the Maillard reaction (MR, also referred to as glycation) affects the structure and function of food proteins. MR induces the conformational and chemical modification of food proteins, not only on the level of IgG/IgE recognition, but also by increasing the interaction and recognition of these modified proteins by antigen-presenting cells (APCs). This affects their biological properties, including digestibility, bioavailability, immunogenicity, and ultimately their allergenicity. APCs possess various receptors that recognize glycation structures, which include receptor for advanced glycation end products (RAGE), scavenger receptors (SRs), galectin-3 and CD36. Through these receptors, glycation structures may influence the recognition, uptake and antigen-processing of food allergens by dendritic cells (DCs) and monocytes. This may lead to enhanced cytokine production and maturation of DCs, and may also induce adaptive immune responses to the antigens/allergens as a result of antigen uptake, processing and presentation to T cells. Here, we aim to review the current literature on the immunogenicity of AGEs originating from food (exogenous or dietary AGEs) in relation to AGEs that are formed within the body (endogenous AGEs), their interactions with receptors present on immune cells, and their effects on the activation of the innate as well as the adaptive immune system. Finally, we review the clinical relevance of AGEs in food allergies

An alternative inhibition method for determining cross-reactive allergens

Inhibition assays are an useful tool to identify the allergen of primary sensitization of cross-reactive allergens. Classical ELISA-based inhibition assays are limited by both the availability of commercial standardized allergen extracts and the experience and knowledge needed for making home-made extracts. Moreover the direct comparison of the inhibition ELISAs outcomes between different laboratories is difficult because of different sources of used allergen extracts and a number of methodological variations. Therefore, we propose a novel ImmunoCap (Phadia, Thermofisher Scientific) based immunoinhibition method with the use of commercially available Caps as the allergen source. The novel ImmunoCap based immunoinhibition method was developed and tested with sera from patients with a well-known cross-reactive sensitization for fig (Ficus carica) and ficus (Ficus benjamina). Results were compared with a classically applied inhibition method, i.e. addition of homemade allergen extract to patient serum. The amount of allergens (fig and ficus extracts) needed to reach a similar degree of inhibition was comparable for both inhibition methods. The ImmunoCap based inhibition assay, in addition to classical inhibition methods, is a valuable tool as the ImmunoCap analyzer and commercial allergens (Caps) are more widely available which makes the outcomes of inhibition tests comparable between different laboratories. Furthermore, in the ImmunoCap inhibition method the same protein source is used for both the inhibition of sIgE and sIgE measurement, which might be even more relevant when multiple cross-reactive allergens are tested

The Indirect Basophil Activation Test Is a Safe, Reliable, and Accessible Tool to Diagnose a Peanut Allergy in Children

Background: The gold standard for the diagnosis of a peanut allergy is an oral food challenge (OFC), but it is a time-consuming, patient-unfriendly, and expensive test. The in vitro direct basophil activation test (BAT) for peanuts was shown to be a promising diagnostic tool for replacing the OFC. Objective: To determine the diagnostic accuracy of the indirect (passive) BAT. Compared with the direct BAT, the timing of the indirect BAT is more flexible, and the problem of nonresponding basophils (unresponsive to IgE receptor–mediated signaling) is circumvented. Methods: In 74 children, suspected of peanut allergy and eligible for an OFC, indirect BAT results for peanut extract, Ara h2, and Ara h6 were compared with the results of a double-blind placebo-controlled food challenge. The reactivity and sensitivity of the basophils in the BAT were correlated to both the allergy status and the threshold dose in the OFC. Results: The combined basophil reactivity for Ara h2 and Ara h6 showed the highest accuracy (94%) for the diagnosis of a peanut allergy, with positive and negative predictive values of 96% and 89%, respectively. The sensitivity of the basophils for Ara h2 significantly discriminates between patients who tolerated up to 0.4 g of peanut protein in the OFC and those who did not. Conclusions: Because the indirect BAT showed a high diagnostic accuracy for peanut allergy, it is a promising alternative to the classical direct BAT and could lead to a reduction in OFC use