a, b. Crry−/−EAMG mice show augmented IFN-γ and IL-4 recall responses to tAChR ex vivo . Splenic cell suspensions (105 cells/well) isolated from WT EAMG and Crry −/− EAMG mice were re-stimulated in triplicate with tAChR at concentrations of 10, 1.0, 0.1, and 0.01 μg/ml. The frequency of IFN-γ and IL-4 secreting cells was determined by ELISPOT assay and additional controls for negative or positive spot production were established. Con A (1 μg/ml) treated cells (positive control) generated comparable number of spots in all experimental groups (243 ± 57 WT and 211 ± 36 Crry −/−), whereas PBS immunized (negative control) mice did not have any spots. Results shown are representative of two independent experiments. Asterisks (*) represent statistically significant differences at p < 0.05.