Figure 4

Western blot analysis using isolated membrane fraction and immunoprecipitates by antibodies in the cultured trophoblasts. (A) The cells were lysed using a teflon/glass homogenizer and a sonicator, and subjected to sucrose gradient centrifugation as described under Materials and Methods. Equal amounts of protein (10 µg) from each of twelve fractions (1-12) were separated by SDS-PAGE 15% acrylamide gel and transferred to membranes, which were blotted using anti-hOAT4 antibody (1:500) and anti-caveolin-1 (1:200). Obtained data from 2 separate experiments were consistent. (B) Cell lysates were immuno-precipitated initially with antibodies of hOAT4 or caveolin-1 and the respective immuno-precipitated (IP) proteins (15 µg) were loaded onto each lane of a 10% SDS-polyacrylamide gel. Cell lysate was immuno-precipitated initially with hOAT4 (left pannel) and was immuno-precipitated initially with caveolin-1 antibodies (right pannel). Obtained data from 2 separate experiments were consistent.