Robust expression of CCR3 as a single basophil selection marker in flow cytometry

Abstract

CCR3 is a stable and highly expressed basophil selection marker, independent of the atopic background or basophil activation state and allows an accurate identification of basophils without need of a second marker. The basophil markers CD123/HLA-DR and IgE showed significantly higher interindividual variability in cell surface expression and are therefore less suited as selection markers.

CCR3 as a single selection marker compared to CD123/HLADR to isolate basophils in flow cytometry: Some comments

“…Another well-known selection approach using the highly basophil specific marker anti-IgE has raised serious objections, because anti-IgE is a potential triggering agent, because other leucocytes express IgE-receptor (Fc‹RII) and because the density of IgE and Fc‹RI may vary widely among subjects and even among basophils in the same individual (3). So, recently published papers have suggested the use of eotaxin receptor CCR3, also known as CD193, as a single selection marker to separate basophils from other leukocytes in flow cytometry, though with some controversial issue (7,8). This marker has the indisputable advantage of being highly expressed in cells at the basal level with a comparable extent in different individuals but it changes its expression upon basophil stimulation (7).…”

“…So, recently published papers have suggested the use of eotaxin receptor CCR3, also known as CD193, as a single selection marker to separate basophils from other leukocytes in flow cytometry, though with some controversial issue (7,8). This marker has the indisputable advantage of being highly expressed in cells at the basal level with a comparable extent in different individuals but it changes its expression upon basophil stimulation (7). Phenotyping markers, which up-or down-regulate membrane expression following cell stimulation, make it very difficult to perform a gating process without introducing cellular contamination into the gate and/or to attain a loss of target cells when activated.…”

α-Galactosylceramide–Induced Airway Eosinophilia Is Mediated through the Activation of NKT Cells

“…CCR3 is highly expressed on eosinophils and a subset of T cells. Human basophils have been reported to express CCR3 in some studies (24); however, there is no CCR3 expression on mouse basophils (25,26). The frequency and absolute number of eosinophils in a-GalCer-administered wild-type mice were significantly increased compared with those in PBS controls.…”

Mast cells and basophils are essential for allergies: mechanisms of allergic inflammation and a proposed procedure for diagnosis

“…Therefore, the identification of a constitutive and constant expression marker for basophils, regardless of activation status, is essential for the specificity of BAT. Thus, CCR3 [119] and CD123 (IL-3 receptor α chain) [120] have been selected as constant expression and identification markers for basophils to distinguish these cells from CD63, the activation marker. Because CCR3 has also been observed on eosinophils [121] and a small proportion of CD4 + T cells [122] , this protein is not the unique marker for basophils.…”

“…However, the IgE gating strategy has the highest variation, showing up to 80% of non-basophils in the selected gate in certain donors [119] . This variation might reflect the large proportion of CD1c + DCs [138] and eosinophils [139] expressing surface FcεRI.…”