Assessing basophil activation by using flow cytometry and mass cytometry in blood stored 24 hours before analysis - 19/04/17
Abstract |
Background |
Basophil activation tests (BATs) have promise for research and for clinical monitoring of patients 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 |
We attempted 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 patients with peanut allergy was collected into 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-IgE or 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 identification of a population of CD63hi basophils, irrespective of whether the specimens were analyzed by means of conventional flow cytometry or cytometry by time-of-flight mass spectrometry, 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 with blood obtained in heparin tubes and stored at 4°C for 24 hours.
Le texte complet de cet article est disponible en PDF.Key words : Basophils, CD63, CD203c, anti-coagulants, heparin, EDTA, peanut allergy, cytometry by time-of-flight mass spectrometry, platelets
Abbreviations used : BAT, CMF-PBS, CyTOF, DAPI, FITC, MFI, OIT, PE, PerCP, RT
Plan
| Supported by National Institutes of Health/National Institute of Allergy and Infectious Diseases grant 5U19AI104209. |
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| Disclosure of potential conflict of interest: S. C. Bendall has received a grant from the National Institutes of Health (NIH) and has consultant arrangements with Fluidigm. H. T. Maecker has received a grant from the NIH (1U19AI104209). R. S. Chinthraja has received a grant and travel support from the National Institute of Allergy and Infectious Diseases. M. Tsai and S. J. Galli have received a grant from the NIH. The rest of the authors declare that they have no relevant conflicts of interest. |
Vol 139 - N° 3
P. 889 - mars 2017 Retour au numéro
Article précédent
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