δ‐Containing GABAA receptors are located extrasynaptically and mediate tonic inhibition. Their involvement in brain physiology positions them as interesting drug targets. There is thus a continued interest in establishing reliable recombinant expression systems for δ‐containing GABAA receptors. Inconveniently, the recombinant expression of especially α4β1/3δ receptors has been found to be notoriously difficult, resulting in mixed receptor populations and/or stoichiometries and differential pharmacology depending on the expression system used. With the aim of developing a facile and robust 96‐well format cell‐based assay for extrasynaptic α4β1/3δ receptors, we have engineered and validated a HEK293 Flp‐In™ cell line stably expressing the human GABAA δ‐subunit. Upon co‐transfection of α4 and β1/3 subunits, at optimized ratios, we have established a well‐defined system for expressing α4β1/3δ receptors and used the fluorescence‐based FLIPR Membrane Potential (FMP) assay to evaluate their pharmacology. Using the known reference compounds GABA and THIP, ternary α4β1/3δ and binary α4β1/3 receptors could be distinguished based on potency and kinetic profiles but not efficacy. As expected, DS2 was able to potentiate only δ‐containing receptors, whereas Zn2+ had an inhibitory effect only at binary receptors. By contrast, the hitherto reported δ‐selective compounds, AA29504 and 3‐OH‐2'MeO6MF, were non‐selective. The expression system was further validated using patch clamp electrophysiology, in which the superagonism of THIP was confirmed. The established FMP assay set‐up, based on transient expression of human α4 and β1/3 subunits into a δ‐subunit stable HEK293 Flp‐In™ cell line, portrays a simple 96‐well format assay as a useful supplement to electrophysiological recordings on δ‐containing GABAA receptors.