Biomaterial Database

Flow cytometric basophil activation tests: Staining of exteriorized basophil granule matrix by fluorescent avidin versus appearance of CD63. 2020

Didier G Ebo, and Jessy Elst, and Michel van Houdt, and Isabel Pintelon, and Jean-Pierre Timmermans, and Tatsuo Horiuchi, and Margaretha A Faber, and Margo M Hagendorens, and Christel M Mertens, and Vito Sabato

Department of Immunology, Allergology, Rheumatology and the Infla-Med Research Consortium of Excellence, University of Antwerp and Antwerp University Hospital, Antwerp, Belgium.

Staining of exteriorized basophil granule matrix by fluorescent avidin might be a reliable technique to monitor basophil degranulation. This study compares the avidin-based technique with the upregulation of CD203c and appearance of CD63 in response to various stimuli. Fourteen individuals responsive to anti-IgE, nine healthy controls, and five birch pollen-allergic patients, and five nonresponders were studied. Activation experiments included anti-IgE, fMLP, interleukin-(IL)-3, and birch pollen allergen. Basophil activation/degranulation was analyzed by flow cytometry and microscopy using anti-CD63, anti-CD203c, and avidin. Stimulation with anti-IgE, fMLP, and relevant allergen results in upregulation of CD203c, CD63 appearance, and an increase in avidin binding. In response to anti-IgE and allergen, upregulation of CD203c peaks within 10 min, CD63 and avidin binding reach a plateau after 10-20 min. CD63 staining leads to a bimodal distribution, avidin staining causes a unimodal shift with a less clear discrimination between degranulating and nondegranulating cells. In response to fMLP, upregulation of CD203c and CD63 and avidin binding are maximal after 2.5 min. Following incubation with anti-IgE and fMLP, percentages of CD203c+ cells are higher than those of CD63+ and avidin+ cells, pointing to a dissociation between activation and degranulation. Percentages of CD63+ cells are systemically higher than those of avidin+ cells. Incubated with IL-3 only upregulates CD203c, while no CD63 or avidin binding is observed. Staining of exteriorized proteoglycans by avidin is a reliable technique to quantify basophil degranulation but offers no added value when compared to traditional assays that use CD63 as a readout.

UI	MeSH Term	Description	Entries
D005434	Flow Cytometry	Technique using an instrument system for making, processing, and displaying one or more measurements on individual cells obtained from a cell suspension. Cells are usually stained with one or more fluorescent dyes specific to cell components of interest, e.g., DNA, and fluorescence of each cell is measured as it rapidly transverses the excitation beam (laser or mercury arc lamp). Fluorescence provides a quantitative measure of various biochemical and biophysical properties of the cell, as well as a basis for cell sorting. Other measurable optical parameters include light absorption and light scattering, the latter being applicable to the measurement of cell size, shape, density, granularity, and stain uptake.	Cytofluorometry, Flow,Cytometry, Flow,Flow Microfluorimetry,Fluorescence-Activated Cell Sorting,Microfluorometry, Flow,Cell Sorting, Fluorescence-Activated,Cell Sortings, Fluorescence-Activated,Cytofluorometries, Flow,Cytometries, Flow,Flow Cytofluorometries,Flow Cytofluorometry,Flow Cytometries,Flow Microfluorometries,Flow Microfluorometry,Fluorescence Activated Cell Sorting,Fluorescence-Activated Cell Sortings,Microfluorimetry, Flow,Microfluorometries, Flow,Sorting, Fluorescence-Activated Cell,Sortings, Fluorescence-Activated Cell
D006801	Humans	Members of the species Homo sapiens.	Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D001360	Avidin	A specific protein in egg albumin that interacts with BIOTIN to render it unavailable to mammals, thereby producing biotin deficiency.
D001491	Basophils	Granular leukocytes characterized by a relatively pale-staining, lobate nucleus and cytoplasm containing coarse dark-staining granules of variable size and stainable by basic dyes.	Basophil
D013194	Staining and Labeling	The marking of biological material with a dye or other reagent for the purpose of identifying and quantitating components of tissues, cells or their extracts.	Histological Labeling,Staining,Histological Labelings,Labeling and Staining,Labeling, Histological,Labelings, Histological,Stainings
D015549	Basophil Degranulation Test	An in vitro test used in the diagnosis of allergies including drug hypersensitivity. The allergen is added to the patient's white blood cells and the subsequent histamine release is measured.	Basophil Activation Test,Activation Test, Basophil,Activation Tests, Basophil,Basophil Activation Tests,Basophil Degranulation Tests,Degranulation Test, Basophil,Degranulation Tests, Basophil,Test, Basophil Activation,Tests, Basophil Activation
D060149	Tetraspanin 30	Ubiquitously-expressed tetraspanin protein that is found in late ENDOSOMES and LYSOSOMES. It functions in intracellular protein transport and signaling.	Antigens, CD63,CD63 Antigen,Granulophysin,LIMP-CD63,Melanoma Antigen ME491,Platelet Glycoprotein 40 kDa,Pltgp40,Antigen ME491, Melanoma,Antigen, CD63,CD63 Antigens