Figure 1

The ITIMs of CD300a are essential for the inhibition of BCR stimulated activation. (A) DT40 chicken B cells expressing CD300a WT or CD300a 4F were loaded with Fluo-4 and Fura-Red. Cells were stimulated with anti-chicken BCR plus isotype control antibody (black line) or anti-chicken BCR plus anti-CD300a mAb (grey line) for 30 seconds and then co-crosslinked with a secondary antibody (GAM). Fluorescence emission was measured in a flow cytometer. Ca²⁺ mobilization is expressed as the ratio of Fluo-4/Fura-Red as a function of time. These results are representative of three independent experiments. (B) DT40 chicken B cells expressing CD300a WT or CD300a 4F were transiently transfected with a NFAT luciferase reporter plasmid and stimulated with GAM plus anti-chicken BCR plus isotype control or anti-chicken BCR plus anti-CD300a mAb. The measured luciferase activity was normalized to the activity obtained with cells treated with PMA plus ionomycin. Data are presented as percentage of inhibition of CD300a vs. isotype control and they are the average ± SEM for three separate experiments.