Immunoblot of claudin-2 expression in clones stably transduced with wild-type (WT) claudin-2 or the following claudin-2 mutants: C54A, C64A, C54A/C64A, C54S, C64S, and C54S/C64S. Cells were cultured in the presence (+) or absence (−) of doxycycline. Cell lysates were subjected to reducing SDS-PAGE and immunoblotted using rabbit claudin-2 antibody. Inducible expression of a band at the predicted molecular mass for claudin-2 monomer (∼20 kDa) was seen in all mutants. Note the appearance of a higher molecular mass band of ∼35 kDa most prominently in C64A and C64S in the absence of doxycycline.