Abstract: A method is disclosed for determining an analyte in a sample suspected of containing the analyte. The analyte is a member of a specific binding pair (sbp) consisting of ligand and complementary receptor. The method comprises combining in an aqueous medium (1) said sample, (2) a first reagent comprising a non-dispersed surface to which is bound an sbp member that becomes bound to the surface in relation to the amount of analyte in the sample. The volume of the aqueous medium containing the above reagents is sufficiently large to allow complete immersion of the first reagent therein and determination of the analyte but not so large to result in substantial interference in the determination upon addition of a second reagent reactive with the conjugate of enzyme and sbp member and capable of generating a signal. The second reagent is present in a medium of sufficient volume to substantially increase the volume of the liquid medium and reduce interference in the determination.