Basophil Activation Test for Allergy Diagnosis

The BAT was performed using heparinized venous blood on the same day of blood collection. Basophils in 100 μL of whole blood were stimulated with the same volume of egg extract (ALK-Abello), baked EW (Sigma-Aldrich, Poole, UK), anti-IgE (1 μg/mL, Sigma-Aldrich, Poole, United Kingdom) or N-formyl-methionyl-leucyl-phenylalanine (f-MLP; 1 μM, Sigma-Aldrich) diluted in Roswell Park Memorial Institute medium (GIBCO, Paisley, United Kingdom) with Roswell Park Memorial Institute alone as negative control. Cells were stained with CD123-FITC, CD203c-PE, human leukocyte antigen (HLA)-DR-PerCP, and CD63-APC (Biolegend, San Diego, Calif) and analyzed by flow cytometry using Fortessa with Diva software (BD Biosciences, San Jose, Calif). Data were analyzed with FlowJo software (version 7.6.1; TreeStar, Ashland, Ore) gating basophils as low side scatter/CD203c+/CD123+/HLA-DR-¹¹ (link)^,¹⁷ (link) cells. Basophil activation was expressed as %CD63+ basophils.

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Marques-Mejias A., Radulovic S., Foong R.X., Bartha I., Krawiec M., Kwok M., Jama Z., Harrison F., Ricci C., Lack G., Du Toit G, & Santos A.F. (2024). Partial Egg Consumption Modifies the Diagnostic Performance of Allergy Tests to Predict Outcome of Double-Blind Placebo-Controlled Food Challenges to Egg. The Journal of Allergy and Clinical Immunology. in Practice, 12(3), 660-669.e5.

Publication 2024

anti-IgE N-formyl-methionyl-leucyl-phenylalanine (f-MLP; CD123-FITC CD203c-PE HLA)-DR-PerCP CD63-APC Fortessa Diva software HLA-DR

Corresponding Organization :

Other organizations : King's College London, Evelina London Children's Healthcare, St Thomas' Hospital, North-West University

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Variable analysis

independent variables

Egg extract (ALK-Abello)
Baked EW (Sigma-Aldrich, Poole, UK)
Anti-IgE (1 μg/mL, Sigma-Aldrich, Poole, United Kingdom)
N-formyl-methionyl-leucyl-phenylalanine (f-MLP; 1 μM, Sigma-Aldrich)

dependent variables

%CD63+ basophils

control variables

Roswell Park Memorial Institute medium (GIBCO, Paisley, United Kingdom)

positive controls

Anti-IgE (1 μg/mL, Sigma-Aldrich, Poole, United Kingdom)
N-formyl-methionyl-leucyl-phenylalanine (f-MLP; 1 μM, Sigma-Aldrich)

negative control

Roswell Park Memorial Institute alone

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