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Figure 3


Fig. 3. (A) PKC{zeta} is present in intestinal sub-cellular fractions as identified by immunoblotting, and antibodies against PKC{zeta} inhibit PCTV budding. Enterocytes were isolated, homogenized and separated into sub-cellular fractions (Materials and Methods). 30 µg protein from each fraction was separated by 12% SDS-PAGE, the proteins transblotted to nitrocellulose membranes and PKC{zeta} identified by anti-PKC{zeta} antibodies using ECL. The different sub-cellular fractions whose proteins are separated on the gel are shown above each lane. (B) The effect of anti-PKC{zeta} antibodies on PCTV-budding activity as a percentage of budding activity using cytosol treated with IgG. Cytosol and ER were treated either with IgG (NC), or anti-PKC{zeta} antibodies (10 µl) (PKC{zeta} Ab) or anti-PKC{zeta} antibodies (10 µl) previously treated with PKC{zeta} antigen (20 µg) (PKC{zeta} Ab+An). Excess antibodies and antigen were removed from the ER by washing, and from the cytosol using anti-IgG bound to beads. Data are the mean ± s.e.m., n=4. P values indicate differences between the mean of PKC{zeta} Ab and either NC or PKC{zeta} Ab+An.