Objective: The voltage-gated potassium channel, Kv7.1 (KCNQ1), is a member of the Kv7 family and is best known for its function in the heart where it contributes to the repolarization of the cardiac action potential. Kv7.1 associates with the regulatory b-subunit KCNE1 and together they underlie the slowly activating delayed rectifier potassium current I_Ks in the heart. Mutations in Kv7.1 and KCNE1 may lead to changes in the duration of the QT-interval and subsequently cardiac arrhythmia. The aim of this study was to elucidate the steady state subcellular localization of Kv7.1 and KCNE1 subunits when expressed alone or together. This was done in order to address how each of the subunits affect the subcellular localization of the Kv7.1/KCNE1 complex and to deduce in which part of the trafficking pathway the complex formation takes place. Methods: MDCK cells were transiently transfected with KCNE1 together with either wild-type (WT) or trafficking deficient Kv7.1 mutants. The subcellular localization of the complex was assessed by confocal microscopy. Results: When singly expressed in MDCK cells, wild type KCNE1 localized to the Golgi apparatus, while WT Kv7.1 was found at the basolateral plasma membrane. Upon co-expression, KCNE1 moved to the cell surface where it co-localized with Kv7.1 in the basolateral plasma membrane. When WT KCNE1 was co-expressed with ER-retained Kv7.1 mutants the localization of KCNE1 was unaffected by the presence of Kv7.1 and KCNE1 was observed in the Golgi complex. In contrast, co-expression of KCNE1 with the endosomally localized 2xL-Kv7.1 mutant resulted in the redistribution of KCNE1 to the same endosomal compartment where it co-localized Kv7.1. Conclusions: Our results indicate that Kv7.1 is the major determinant of the subcellular localization of the Kv7.1/KCNE1 complex. Furthermore, our data suggest that stable assembly of Kv7.1 and KCNE1 takes place within the Golgi apparatus.