Multiple classes of pharmacological agents including benzodiazepines, cage convulsants like t-butylbicyclophosphorothionate (TBPS), barbiturates and neuroactive steroids allosterically modulate the γ-aminobutyric acid A receptor-chloride ionophore complex (GRC). The function of benzodiazepines requires a GRC comprised of α, β and γ subunits, while TBPS, barbiturates and neuroactive steroids will allosterically modulate GRCs comprised of only α and β subunits. Binary αβ complexes are still hypothesized to be expressed in the mammalian brain particularly during development and could contribute to the pharmacological action of neuroactive steroids and barbiturates. In order to examine binary αβ complexes we report here the establishment of stable cell lines that express high levels of human GABA A receptors comprised of α1β1, α2β1 and α3β1 subunit combinations. The apparent potencies for allosteric modulation of [ 3 5 S]TBPS for most naturally occurring neuroactive steroids for the binary subunit combinations was similar to that of the γ-containing subunit combinations. Also discussed is the usefulness of these cell lines for the biophysical analysis of the GABA A receptor stoichiometry.