Test: Basophil Activation Test - Ara h 2

Video Link

Background/Purpose

The basophil activation test (BAT) is a functional assay that measures the degree of degranulation following stimulation with allergen or controls by flow cytometry. It correlates directly with histamine release. From the dose-response curve resulting from BAT in allergic patients, basophil reactivity (%CD63+ basophils) and basophil sensitivity (EC50 or similar) are the main outcomes of the test. BAT takes into account all characteristics of IgE and allergen and thus can be more specific than sensitization tests in the diagnosis of allergic disease. BAT reduces the need for in vivo procedures, such as intradermal tests and allergen challenges, which can cause allergic reactions of unpredictable severity.

Procedure

Blood is collected in an EDTA-containing Vacutainer tube and mixed gently. Stimulation and lysing buffers are prewarmed to room temperature.
Allergens are diluted following a specific protocol.
Allergenes are mixed with the blood according to specific protocols.
Staining reagent containing anti-CCR3-PE and anti-CD63-FITC are added subsequently.
All tubes are mixed, covered, and incubated at 37°C for 25 minutes in an incubator, after which 2 mL of lysing reagent is added and each tube vortexed gently and incubated for 10 minutes at room temperature in the dark.
After centrifugation at 500 × g for 5 minutes, supernatants are decanted gently and pellets resuspended and kept at 4°C until analyses.
Flow cytometry is performed at each site and data is analysed using software
Basophils are gated as SSClow/CCR3+ and activation is expressed as %CD63+ basophils, corrected for the spontaneous basophil activation (ie, subtracted the %CD63+ basophils in the unstimulated condition).
Performed: Bilaterally

Dialog

Blood based analysis of BAT test using CD63 activation.

Criteria

• Basophil activation level below threshold.

Measurement

%

LOINC Code

Scale/Grading

selection range; grade - LOINC ID
Option - Value option selection range; grade - LOINC IDPass % ; Pass - Fail % ; Fail -