Fig. 3. Effect of CAL on m2apKO T. gondii. (A) Western blotting of microneme proteins released into supernatants indicated that m2apKO parasites were unable to secrete MIC2 even after CAL treatment, unlike wild-type parasites (RH strain). Stimulation with 1% ethanol was used as a positive control for secretion. Supernatants were compared to diluted lysates of cell standards (% pellets). Identical blots were probed with monoclonal antibody 6D10 (cellular MIC2 and secreted MIC2; upper panel) and rabbit polyclonal serum to MIC5 (lower panel). (B) Phosphorimager analysis of western blots demonstrated that the amount of MIC2 secretion by the m2apKO strain was less than 10% of wild-type (RH strain) secretion even after CAL treatment. However, secretion of the microneme protein MIC5 was normal. Bars represent the average of three experiments (mean±s.e.m.). (C) Quantification of SAG1-labeled trails deposited in gliding assays demonstrated that average trail length formed by the m2apKO strain did not increase with CAL treatment, unlike trail length in wild-type parasites (RH). 1 µM cytochalasin D (CD) was used as a negative control for gliding. Bars show average trail length in parasite body lengths (mean±s.e.m.). ^*A significant difference when compared to control trail lengths; P0.05, paired Student's t-test.