HEK293 cells were transiently transfected with the 4 estrogen-responsive elements containing vector together and an alligator ESR expression vector (A and B) or were transfected with pG5luc vector containing 5 GAL4 binding sites and LBD of each ESR in pBIND vector, including the GAL4 DNA-binding domain (C and D). Cells were incubated with serial concentrations of E2, PPT, DPN, FERb 033, or WAY 200070 at 10−14M–10−5M. Data were represented as fold inductions of luciferase signals as compared with vehicle (DMSO) controls. PPT and WAY 200070 showed the highest specificity on ESR1 and ESR2, respectively. Each column represents the mean of triplicate determinations, and vertical bars represent the mean ± 1 SEM.