The Generation and Characterization of Recombinant Protein and Antibodies of Clostridium perfringens Beta2 Toxin

Figure 7

Ability of rCPB2 binding to NCM460 cells characterized by an immunofluorescent assay. NCM460 cells were incubated with rCBP2 at concentration of 15 μg/mL for indicated times. The cells were then used for accessing rCPB2 protein and cell membrane with McAb 1E23 to CPB2 (1 : 100) and rabbit polyclonal antibody to Annexin A2 (1 : 200), respectively. FITC-labeled donkey anti-mouse IgG (1 : 500) and Rhodamine- (TRITC-) conjugated goat anti-rabbit IgG (1 : 250) were used as secondary antibodies for visualizing respective proteins of interest. DAPI was used for nuclear staining. (a–f) Representative images of immunofluorescent staining for cells incubated with rCPB2 at indicated times of 5 min (a), 30 min (b), 2 h (c), 4 h (d), 6 h (e), and 12 h (f). The results showed that rCPB2 was able to bind to the membrane of NCM460 cell and translocate into the cytoplasm of cells. Insets were the enlarged boxed regions of respective images in (a–f). Bars in (a–f): 50 μm.

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