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Assessing basophil activation by flow cytometry and mass cytometry in blood stored 24 hours before analysis #MMPMID27527263
J Allergy Clin Immunol 2017[Mar]; 139 (3): 889-899.e11 PMID27527263show ga
Background: Basophil activation tests (BATs) have promise for research and for clinical monitoring of subjects with allergies. However, BAT protocols vary in blood anticoagulant used and temperature and time of storage before testing, complicating comparisons of results from various studies. Objective: To attempt to establish a BAT protocol that would permit analysis of blood within 24 hours of obtaining the sample. Methods: Blood from 46 healthy donors and 120 peanut allergic patients was collected into ethylenediaminetetraacetic acid (EDTA) or heparin tubes, and samples were stored at 4°C or room temperature for 4 or 24 hours before performing BATs. Results: Stimulation with anti-Immunoglobulin E (anti-IgE) or interleukin-3 (IL-3) resulted in strong upregulation of basophil CD203c in samples collected in EDTA or heparin, stored at 4°C, and analyzed 24 hours after sample collection. However, a CD63hi population of basophils was not observed in any conditions in EDTA-treated samples unless exogenous calcium/magnesium was added at the time of anti-IgE stimulation. By contrast, blood samples collected in heparin tubes were adequate for quantification of upregulation of basophil CD203c and for identification of a population of CD63hi basophils, whether the specimens were analyzed by conventional flow cytometry or by Cytometry by Time-of-Flight mass spectrometry (CyTOF), and such tests could be performed after blood was stored for 24 hours at 4°C. Conclusion: BATs to measure upregulation of basophil CD203c and induction of a CD63hi basophil population can be conducted using blood obtained in heparin tubes and stored at 4°C for 24 hours.