C. H. Bridts

Flow-assisted allergy diagnosis: current applications and future perspectives

Physicians predominantly rely upon quantification of serum‐specific immunoglobulin E (IgE) and/or skin test to confirm clinically suspected IgE‐mediated allergy. However, for various reasons, identification of the offending allergen(s) and potentially cross‐reactive structures is not always straightforward. Flow‐assisted allergy diagnosis relies upon quantification of alterations in the expression of particular basophilic activation markers. Actually, upon challenge with a specific allergen, basophils not only secrete quantifiable bioactive mediators but also upregulate the expression of different markers which can be detected efficiently by flow cytometry using specific monoclonal antibodies. Currently, the technique has been applied in the investigation of IgE‐mediated allergy caused by classical inhalant allergens, food, Hevea latex, hymenoptera venoms and drugs. It is also appreciated; the technique proves valuable in the diagnosis of non‐IgE‐mediated (anaphylactoid) reactions such drug hypersensitivity and the detection of autoantibodies in certain forms of chronic urticaria. This review will not address immunologic features, characteristics and general pitfalls of flow‐assisted analysis of in vitro‐activated basophils as summarized elsewhere. After a recapitulation of the principles and some specific technical issues of flow‐assisted analysis of in vitro‐activated basophils, we principally focus on the current clinical and research applications of the basophil activation tests. Personal experience of both research groups is provided, where appropriate. Finally, a viewpoint on how the field might evolve in the following years is provided.

Anaphylaxis during anaesthesia : diagnostic approach

Correct management of anaphylaxis during anaesthesia requires a multidisciplinary approach with prompt recognition and treatment of the acute event by the attending anaesthesiologist, and subsequent determination of the responsible agent(s) with strict avoidance of subsequent administration of all incriminated and/or cross‐reacting compounds.

Basophil activation test by flow cytometry: Present and future applications in allergology†

The diagnosis of allergic reactions in clinical practice rests upon both clinical history and the demonstration of specific immunoglobulin E (sIgE), either in the serum or via skin tests. However, for various reasons, identification of the offending allergen(s) is not always possible. Moreover, not all allergies are IgE‐mediated. In an attempt to find reliable methods to investigate hypersensitivity reactions, histamine and sulfidoleukotriene release tests have long been introduced. However, relatively few comprehensive quality reports have been published so far. Upon challenge with a specific allergen, basophils not only secrete quantifiable bioactive mediators but also upregulate the expression of different markers which can be detected efficiently by flow cytometry using specific monoclonal antibodies. This review addresses the principals, particular technical aspects and pitfalls as well as the clinical and research applications of flow‐assisted analysis of in vitro activated basophils

In vitro allergy diagnosis: should we follow the flow?

During the last 5 years, an increasing number of studies have demonstrated that flow cytometric quantification of in vitro basophil activation can be a quite performant and reliable tool to measure IgE‐dependent allergen‐specific responses in allergic patients. So far, most assays have used CD63 as a basophil activation marker and native allergen extracts for stimulation. However, other basophil markers and recombinant allergens have recently been introduced. The technique has been applied for the diagnosis of allergy to pollen, house dust mite, food, natural rubber latex, hymenoptera venom and drugs. In addition, the technique has proven to be useful in non‐IgE‐mediated reactions such as hypersensitivity to drugs as well as detection of auto‐antibodies in chronic urticaria. This review will focus on some specific issues: (1) principles of flow cytometric analysis of in vitro‐activated basophils, (2) general technical aspects of the technique (including passive sensitization), (3) clinical applications and (4) recommendations for further development and evaluation of the technique.

Sensitization to cross-reactive carbohydrate determinants and the ubiquitous protein profilin: mimickers of allergy

Background During the last decade, evidence has been provided for profilins and cross‐reactive carbohydrate determinants (CCDs) to be capable of inducing cross‐reactive IgE antibodies with little clinical relevance.

Influence of pro-inflammatory (IL-1α, IL-6, TNF-α, IFN-γ) and anti-inflammatory (IL-4) cytokines on chondrocyte function

Abstract Objective: Cytokines produced by inflammatory cells play a pivotal role in synovial inflammation and joint destruction in rheumatoid arthritis. To investigate the influence of pro-inflammatory cytokines (IL-1α, IL-6, TNF-α, IFN-γ) and subsequently the possible beneficial role of an anti-inflammatory cytokine (IL-4) on chondrocyte viability (necrosis/apoptosis), proliferation and nitric oxide (NO) production. Methods: Primary bovine chondrocytes were cultured until monolayers were obtained. Cells were incubated with cytokines (IL-1α, IFN-γ, TNF-α, IL-4, IL-6) at 0.1, 1, 10 and 100ng/mL. After 48h, the viability of the chondrocytes was measured flow cytometrically with propidium iodide. Proliferation was determined by the incorporation of tritiated thymidine. The morphology of the chondrocytes, including presence of apoptotic nuclei, was evaluated by a May-Grunwald–Giemsa staining. In addition, the number of apoptotic chondrocytes was detected flow cytometrically with a TUNEL technique and annexin-V/propidium iodide staining. NO production was evaluated using a spectrophotometric assay, based upon the Griess reaction. Results: The viability and proliferation of bovine chondrocytes decreased after incubation with 100ng/mL IL-1α, TNF-α or IFN-γ. In contrast, incubation of chondrocytes with IL-4 or IL-6 had no influence on the viability or the proliferation of cells. IL-1α was able to enhance NO production in a dose dependent manner. IFN-γ and TNF-α induced NO production only at the highest concentration (100ng/mL), whereas IL-4 and IL-6 did not. There was a dose dependent increase in apoptosis of bovine chondrocytes cultured in the presence of IL-1α and TNF-α. This effect could not be prevented by preincubation with IL-4. Preincubation with IL-4 diminished IL-1α and TNF-α induced NO production and increased proliferation of chondrocytes. In an additional experiment, incubation of human chondrocytes with anti-Fas did not induce apoptosis as measured by annexin-V/propidium iodide staining. Conclusions: Pro-inflammatory cytokines are able to induce apoptosis, whereas IL-4 as an anti-inflammatory cytokine can inhibit the effect of IL-1α and TNF-α on NO production and proliferation of bovine chondrocytes.

Differences in circulating dendritic cell subtypes in cord blood and peripheral blood of healthy and allergic children

Background Different types of circulating dendritic cells have been described. Dendritic cells influence differentiation of naive T lymphocytes into T helper type 1 (Th1) and Th2 effector cells.

Component‐resolved diagnosis from latex allergy by microarray

Background A positive specific IgE (sIgE) result for latex does not always mirror the clinical situation and is frequently found in individuals without overt latex allergy.

Flow-Assisted Quantification of In Vitro Activated Basophils in the Diagnosis of Wasp Venom Allergy and Follow-up of Wasp Venom Immunotherapy

Correct identification of the culprit venom is a prerequisite for specific venom immunotherapy (VIT). Despite the efficacy of VIT, issues as how to monitor treatment and when to discontinue maintenance therapy remain to be established.

Flow-assisted diagnostic management of anaphylaxis from rocuronium bromide

Background:  Diagnosis of anaphylaxis from neuromuscular blocking agents (NMBA) is not always straightforward.