These tools will no longer be maintained as of December 31, 2024. Archived website can be found here. PubMed4Hh GitHub repository can be found here. Contact NLM Customer Service if you have questions.


Pubmed for Handhelds

PUBMED FOR HANDHELDS

Search MEDLINE/PubMed

  • Title: Analyzing histamine release by flow cytometry (HistaFlow): a novel instrument to study the degranulation patterns of basophils.
    Author: Ebo DG, Bridts CH, Mertens CH, Hagendorens MM, Stevens WJ, De Clerck LS.
    Journal: J Immunol Methods; 2012 Jan 31; 375(1-2):30-8. PubMed ID: 21945198.
    Abstract:
    BACKGROUND: Stimulated human basophils exhibit different degranulation patterns with release of mediators and appearance of activation markers such as CD63 and CD203c. Traditionally, released mediators are quantified in the supernatant of activated cells, whereas the expression of activation markers by individual cells is analyzed by flow cytometry. Alternatively, intracellular histamine and its release by basophils and mast cells have been repeatedly studied applying an enzyme-affinity-gold method based on the affinity of the histaminase diamine oxidase for its substrate histamine. OBJECTIVE: To develop a flow cytometric technique enabling to study histamine release by individual basophils in combination with the expression of activation markers. To elucidate the principles of basophil degranulation on a single cell level. METHODS: Intracellular histamine and its release is analyzed flow cytometrically by an enzyme-affinity method using diamine oxidase conjugated to laser-excitable fluorochromes. Phenotyping of cells implied flow cytometric quantification of CD63 and CD203c. Stimuli such as allergen, anti-IgE, N-formyl-met-leu-phe (fMLP), phorbol 12-myristate 13-acetate (PMA), ionomycin and interleukin (IL-)3 are applied to obtain different degranulation profiles. RESULTS: Stimulation with anti-IgE, allergen, fMLP and PMA±ionomycin induces a rapid release of histamine that can be analyzed flow cytometrically. Analyses on a single cell level reveal that histamine release is not restricted to cells showing significant up-regulation of CD63. Alternatively, up-regulation of CD203c does not per se indicate histamine release. In some patients, priming of cells with IL-3 not only facilitates basophil responsiveness but also implies an increased ability of DAO to label the cells. CONCLUSION: This study provides the proof-of-concept that histamine and its release can be studied by multicolor flow cytometry on a single cell level (HistaFlow). Coupling the data to simultaneous phenotyping of activated basophils confirms that histamine release principally results from anaphylactic degranulation and in a lesser extent from piecemeal degranulation.
    [Abstract] [Full Text] [Related] [New Search]