Mouse c kit clone 2b8

1 × 10⁶ cells in 0.1 ml of HNA were incubated with 2.4G2 or human IgG for 15 minutes on ice, then stained with 1 μg each of fluorochrome-labeled mAbs specific for mouse IgE (clone RME-1, Biolegend), human IgE (clone MHE-18, Biolegend), or the anti-huFcεRIα mAbs AER-37 or 15.1, and/or mAbs to human c-kit (clone 104D2, Biolegend) mouse c-kit (clone 2B8, Biolegend), and/or human IL-3R (clone 6H6, Biolegend), mouse B220 (clone RA3–6B2, Biolegend) and/or human CD3 (clone OKT3, Biolegend), or mouse CD3 (clone 145–2C11). Cells were washed twice, fixed with 2% paraformaldehyde, and analyzed with a Becton-Dickinson LSR II Flow Cytometer (BD Biosciences). Mast cells were identified as high forward and side scatter cells that expressed IgE, FcεRIα, and Kit. Basophils had lower side and forward scatter and expressed IL-3R, IgE, and FcεRIα, but lacked Kit.

1 × 10⁶ cells in 0.1 ml of Hank’s balanced salt solution with 10% newborn calf serum and 0.2% sodium azide (HNA) were incubated with 1 μg of 2.4G2 or human IgG for 15 minutes on ice, then stained with 1 μg each of fluorochrome-labeled mAbs specific for mouse IgE (clone RME-1, Biolegend), hu IgE (clone MHE-18, Biolegend), or the anti-huFcεRIα mAbs AER-37 or 15.1, and/or mAbs to human c-kit (clone 104D2, Biolegend) or mouse c-kit (clone 2B8, Biolegend), and human IL-3 receptor (clone 6H6, Biolegend), or mouse IL-3 receptor (clone 5B11, Biolegend), mouse B220 (clone RA3-6B2, Biolegend) and/or human CD3 (clone OKT3, Biolegend), or mouse CD3 (clone 145-2C11) or mouse pan-NK cells marker CD49b (clone DX5, Biolegend); Syk (clone 4D10.2, Biolegend); phospho-Syk (clone moch1ct, ThermoFisher); phospho-SHIP (clone P1C1-A5, Biolegend) Akt (C67E7, from Cell Signaling Tech.), phospho-AKT (D9E, Cell Signaling Tech.), phospho-ERK (clone 4B11B69, Biolegend). Cells were washed twice, fixed with 2% paraformaldehyde, and analyzed with a Becton-Dickinson LSR II Flow Cytometer (BD Biosciences). Mast cells were identified as high side scatter, c-kit positive, IL-3 receptor positive, CD3 negative and B220 negative cells. Basophils were identified as forward/side scatter intermediate, IL-3 receptor positive, CD49b positive, CD3 negative cells^{16 (link)}.