Figure 9. Protein was extracted from native or cultured RPE. Native RPE was isolated from E7, E10, E14, and E18 embryos. For cultured RPE, RPE isolated from E7 or E14 embryos was cultured with SF2 in the basal medium chamber and either SF3 or rcSF3 in the apical medium chamber. The proteins were resolved by electrophoresis, blotted onto membranes and probed with antibodies to the indicated protein. Antibodies directed against claudin 19 were not available. In preliminary experiments, proteins of the appropriate M_r were detected, and the corresponding peptide antigen could compete for binding of the antibody (data not shown). As inferred from mRNA levels, the relative amounts of different claudin family members varied among the different stages of development. The effects of rcSF3 on claudin expression were variable and, in some instances, was not predicted by changes in mRNA levels. The discrepancies between steady-state levels of mRNA and protein were especially large for ZO-3. The data are representative of 3 independent experiments.