Luc S. De Clerck

Basophil activation tests: time for a reconsideration

Challenges in in vitro allergy diagnostics lie in the development of accessible and reliable assays allowing identification of all offending allergens and cross-reactive structures. Flow-assisted analysis and quantification of in vitro activated basophils serves as a diagnostic instrument with increasing applications developed over the years. From the earliest days it was clear that the test could constitute a diagnostic asset in basophil-mediated hypersensitivity. However, utility of the basophil activation test should be reassessed regarding difficulties with preparation, characterization and validation of allergen extracts; availability and the potential of more accessible diagnostics. Today, the added value mainly lies in diagnosis of immediate drug hypersensitivity. Other potential indications are monitoring venom-immunotherapy and follow-up of natural history of food allergies. However, results in these nondiagnostic applications are preliminary. We review the most relevant clinical applications of the basophil activation test. Some personal comments and views about perspectives and challenges about flow-assisted allergy diagnosis are made.

Moxifloxacin hypersensitivity: Uselessness of skin testing

Quinolones are synthetic antibiotics based on the 4-quinolone and 1,8-naphthyridine nuclei. They can cause severe hypersensitivity reactions, even after first exposure, with moxifloxacin being an important culprit. Unfortunately, a diagnosis of quinolone hypersensitivity is not straightforward, because of the absence of drug-specific sIgE immunoassays and uncertainties associated with skin testing with potentially nonspecific histamine releasers. These contradictory studies show that establishing nonirritating skin-test concentrations for ciprofloxacin is not straightforward. For example, Broz et al recommend an intradermal test (IDT) concentration of 1:300 to 1:1000 in saline for ciprofloxacin. In contrast, the Empedrad study failed to determine a nonirritating concentration for ciprofloxacin because the irritating concentration varied a 1000-fold in healthy volunteers. Accordingly, the European Network on Drug Allergy and European Academy of Allergy and Clinical ImmunologyDrugAllergy InterestGrouphasmadenodrugspecific recommendations about skin prick test (SPT) and/or IDT with quinolones. For moxifloxacin, the literature reveals “nonirritant” concentrations for SPT to vary between 1 and 20 mg/mL and for IDT between 0.004 and 0.05 mg/mL. However, inspection of the data discloses that 3/5 studies did not include controls and data gathered in patients were restricted to a maximum of 5 cases. This study assessed SPT and IDT in the diagnosis of immediate moxifloxacin hypersensitivity. A comparative study was performed between patients with histories of reactions and controls who tolerated a graded oral drug provocation test (DPT) with moxifloxacin. Patient inclusion was based on clinical history. Fourteen patients with a history of an immediate hypersensitivity reaction to moxifloxacin were enrolled. All patients suffered from urticaria/angioedema, bronchospasm/hypotension, and/or loss of consciousness within 60 minutes after intake. Other possible causes were excluded (ie, concomitant intake of other drugs). Time between reaction and investigation ranged between 0.3 and 21 months (median: 4 months) (Table I). Sixteen individuals who tolerated a graded oral DPT served as an exposed-control group. Skin testing consisted of SPTs and IDTs with a parenteral formulation ofmoxifloxacin hydrochloride (Avelox 400mg/250mL [or 1.6 mg/mL], Bayer SA-NV, Diegem, Belgium). SPT was performed using a 1:10 dilution and undilutedmoxifloxacin. Results were considered positive when wheal/flare equaled or exceeded 3/3 mm. For IDTs, serial dilutions of 1:1000, 1:100, and 1:10 were used. Results were considered positive when the wheal equaled or exceeded 5 mm. Patients with a history of betalactam hypersensitivity are more prone to develop hypersensitivity reactions to fluoroquinolones. Because the majority of subjects included as controls consulted with such a history, we preferred a graded DPT instead of a full dose provocation in search of a save alternative. All controls had a negative oral graded DPT with moxifloxacin (Avelox) up to a cumulative dose of 750 mg. Statistical analysis was performed with IBM SPSS Statistics 22 (IBM Corp, Armonk, New York). Fisher’s exact tests were used to evaluate significance differences of the skin-test results between patients and controls. SPT and IDT results are displayed in Figure 1. SPT was negative in all patients and controls. Intradermal testing yielded positivity in 10/14 patients. Two patients tested positive at the 1:1000 dilution (0.0016 mg/mL), 2 patients at the 1:100 dilution (0.016 mg/mL), and 6 patients at the 1:10 dilution (0.16 mg/ mL). Of the 14 patients, 4 displayed a negative SPT and IDT. In controls, IDT was also positive in 12 individuals. Of the 16 individuals, 2 tested positive at the 1:100 dilution. Of the 14 individuals who received the 1:10 dilution, 12 tested positive. The remaining 2 manifested a (true) negative result in both SPT and IDT. Sensitivity and specificity of the intradermal testing calculated between patients and controls in this population were 57% and 12.5%, respectively. Positive and negative predictive values were 36% and 25%, respectively. A comparison of IDT between patients and controls yieldedP-values of .192 for the 1:1000 dilution, 1.00 for the 1:100 dilution, and .209 for the 1:10 dilution. This is the first assessment of skin testing for moxifloxacin with a comparative study between patients and controls who tolerated a cumulative dose of 750 mg moxifloxacine during a graded oral DPT. We conclude that SPTs and IDTs with moxifloxacin hydrochloride are unreliable methods to establish a diagnosis of immediate moxifloxacin hypersensitivity. Our data are in line with previous findings that quinolones frequently induce skin-test responses in healthy subjects. It appears that at higher dilutions skin tests remain negative in a majority of patients, whereas higher concentrations are not discriminative between patients and exposed controls. There are studies that show that the time elapsed between the acute reaction and skin tests affects the outcome of the tests, which makes it difficult to interpret. We were unable to assess such an effect because the majority of patients received skin testing within 6 months after the acute reaction, and a longitudinal follow-up study does not seem to be warranted. A limitation of our study is the absence of controlled DPTs as previous research has shown that as few as 1/3 of patients with similar recent convincing reaction histories to quinolones reacted on being challenged with the culprit drug. However, because the time between the reactions and evaluation was relatively short

Influence of IL‐6, IL‐33, and TNF‐α on Human Mast Cell Activation: Lessons from Single Cell Analysis by Flow Cytometry

Mechanisms that govern priming and degranulation of human mast cells (MCs) remain elusive. Besides, most of our knowledge is based on experiments from which data only reflect an average of all stimulated cells. This study aims at investigating the effects of proinflammatory cytokines IL‐6, IL‐33, and TNF‐α on IgE‐dependent and IgE‐independent activation of individual MCs.