Analyzing Basophil Activation in the Presence of a Test Allergen using Flow Cytometry

Published: November 30, 2023

Abstract

Source: Doña, I. et al., Basophil Activation Test for Allergy Diagnosis. J. Vis. Exp. (2021)

In this video, we demonstrate the test to evaluate basophil activation in the presence of a test allergen using flow cytometry.

Protocol

All procedures involving sample collection have been performed in accordance with the institute's IRB guidelines. 1. Sample preparation Collect peripheral blood in 9 mL heparinized tubes and maintain the sample at room temperature (RT) in a rotor until it is required for the experimental protocol. Label 5 mL cytometer tubes for negative controls (2 tubes), positive controls (2 tubes), and different concentrations of allergen/drug (1 t…

Divulgazioni

The authors have nothing to disclose.

Materials

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

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Citazione di questo articolo
Analyzing Basophil Activation in the Presence of a Test Allergen using Flow Cytometry. J. Vis. Exp. (Pending Publication), e21839, doi: (2023).

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