Figure 6. Effect of peptide concentration on the ER release of calcium.

COS-7 cells expressing MHC class II molecules I-A^K were loaded overnight with increasing concentrations of HEL 46–61 peptides. Calcium responses in 3A9 hybridomas with 2-APB were analyzed with MAAACS. (A) Percentage of responding cells for various peptide concentrations was obtained with activation thresholds (TH_act) respectively for hybridomas stimulated with increasing doses of 1.4 (0.5 nM), 1.8 (5 nM), 1.7 (50 nM), 1.6 (500 nM), 1.5 (5 µM) with 2-APB. (B) Fluorescence amplitude of calcium response of activated cells as a function of peptide concentration is represented on a scatter plot. (C) Response fraction of activated cells as a function of peptide concentration is represented on a scatter plot. (D) Frequency calcium response bursts of activated cells as a function of peptide concentration is represented on a scatter plot. The mean value +/− SEM is represented in red. Statistical tests were carried out using Mann Whitney's non-parametric test (*** = p<0.001; ** = 0.001<p<0.01; * = 0.01<p<0.05; ns = p>0.) (E) Mode of calcium response: For each antigenic peptide concentration, modes of calcium response are represented as a pie chart: unactivated in gray, maintained in pink, oscillating in green and unique in yellow.