Basophil Activation Test for Allergy Diagnosis

Summary

The basophil activation test is a complementary in vitro diagnostic test for the evaluation of IgE-mediated allergic reactions based on the detection of basophil activation in the presence of a specific stimulus through the measure of activation markers by flow cytometry.

Abstract

The basophil activation test (BAT) is a complementary in vitro diagnostic test that can be used in addition to clinical history, skin test (ST), and specific IgE (sIgE) determination in the evaluation of IgE-mediated allergic reactions to food, insect venom, drugs, as well as some forms of chronic urticaria. However, the role of this technique in the diagnostic algorithms is highly variable and not well determined.

BAT is based on the determination of basophil response to allergen/drug cross-linking IgE activation through the measurement of activation markers (such as CD63, CD203c) by flow cytometry. This test can be a useful and complementary tool to avoid controlled challenge tests to confirm allergy diagnosis, especially in subjects experiencing severe life-threatening reactions. In general, the performance of BAT should be considered if i) the allergen/drug produces false positive results in ST; ii) there is no allergen/drug source to use for ST or sIgE determination; iii) there is discordance between patient history and ST or sIgE determination; iv) symptoms suggest that ST may result in systemic response; v) before considering a CCT to confirm the culprit allergen/drug. The main limitations of the test are related to non-optimal sensitivity, especially in drug allergy, the need to perform the test no longer than 24 h after sample extraction, and the lack of standardization between laboratories in terms of procedures, concentrations, and cell markers.

Introduction

IgE-mediated allergy diagnosis is based on clinical history, skin tests (STs), quantification of serum specific IgE (sIgE), and, if it is required and indicated, controlled challenge tests (CCTs)^1,2,3,4,5,6. However, clinical history can be unreliable since there may be a lack of accurate information, and STs and CCTs are not risk-free procedures that can be contraindicated in subjects experiencing severe life-threatening reactions^1,

Protocol

The protocol performance was conducted according to the Declaration of Helsinki principles and approved by the local Ethics Committee (Comité de Ética para la Investigación Provincial de Málaga, Spain). All subjects were informed orally about the research study and they signed the corresponding informed consent form.

NOTE: The present protocol details the BAT procedure that the authors use daily. However, this is not a standardized method and differences with proce.......

Discussion

BAT is a complementary in vitro diagnostic test for the evaluation of IgE-mediated allergic reactions that has shown to be useful in the diagnosis of reactions induced by different triggers such as drugs, food, or inhalants, as well as in some forms of chronic urticaria. In general, BAT performance should be considered if i) the allergen/drug produces false positive results in ST; ii) the allergen/drug is not available to be used for ST or sIgE quantification; iii) discordance between clinical history and ST or .......

Materials

Name	Company	Catalog Number	Comments
5 mL Round Bottom Polystyrene Test Tube, without Cap, Nonsterile	Corning	352008
APC anti-human CD193 (CCR3) Antibody	BioLegend	310708
BD FACSCalibur Flow Cytometer	BD Biosciences
Calcium chloride	Sigma-Aldrich	C1016
FITC anti-human CD63 Antibody	BioLegend	353006
HEPES (1 M)	Thermo-Fisher	15630106
Lysing Solution 10x concentrated	BD Biosciences	349202
Magnesium chloride	Sigma-Aldrich	M8266
N-Formyl-Met-Leu-Phe	Sigma-Aldrich	F3506
PE anti-human CD203c (E-NPP3) Antibody	BioLegend	324606
Potassium chloride	Sigma-Aldrich	P9541
Purified Mouse Anti-Human IgE	BD Biosciences	555857
Recombinant Human IL-3	R&D Systems	203-IL
Sheath Fluid	BD Biosciences	342003
Sodium chloride	Sigma-Aldrich	S3014
TUBE 9 mL LH Lithium Heparin	Greiner Bio-One	455084
Tween 20	Sigma-Aldrich	P1379