FIG. 2.

Determination of integrity of the ER. (A) Cells were hypotonically permeabilized and subjected to proteinase K digestion before (lane 1) and after (lanes 2 to 5) the addition of Triton X-100. Lane 6 shows the total BiP released after treatment with detergent. After treatment, proteins were precipitated and analyzed by 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis, transferred to nitrocellulose, and probed with specific monoclonal antibodies raised against the ER BiP protein. (B) Lanes 1 to 3, assays with permeabilized tachyzoites; lane 4, mock assay with permeabilized tachyzoites under the conditions used for the labeling procedure (37°C incubation); lane 5, permeabilized tachyzoites that were mock labeled and treated with 3 U/ml B. thuringiensis PI-PLC. Each lane was loaded with a cell equivalent of 5 × 10⁷ tachyzoites.