Neutralization of CT holotoxin by pCTB-induced antibodies in the CHO cell elongation assay.

A and B, quantitation of the CHO cell elongation by CT holotoxin pre-treatment with 10× diluted (A) fecal and (B) serum Ig samples. The holotoxin (20 ng/ml) was pre-incubated with 10× diluted fecal or serum samples obtained 2 weeks after the second vaccination from animals immunized with PBS, 10 or 30 µg pCTB, or 30 µg eCTB. Cell length was measured after overnight incubation and crystal violet staining. See Experimental Procedure for the detailed method. Data are shown as mean ± SEM of triplicate. For both fecal and serum Igs, all immunized groups showed significant neutralization of CT-induced cell elongation compared to the PBS control group: ***P<0.001 (ANOVA with Bonferroni's multiple comparison test). C, cells treated with CT. D–G, cells treated with CT and fecal samples from animals immunized with 10 µg pCTB, 30 µg eCTB, 30 µg pCTB, and PBS, respectively. H, untreated control cells. I–L, cells treated with CT and serum samples from animals immunized with 10 µg pCTB, 30 µg eCTB, 30 µg pCTB, and PBS, respectively. Scale bar, 100 µm.