There is described an assay for determining the relative concentrations of two antigenic solutes in a sample. The assay involves a first antibody to the first antigenic solute, a second antibody to the second antigenic solute and a ligand molecule having substituents to which the first and second antibodies may bind. The second antibody is immobilized upon a solid phase support and the other components are in solution. The components are mixed with a sample containing the antigenic solutes causing competition between the ligand molecule and each of the antigenic solutes for binding to the first and second antibodies. In the situation where there is a relatively higher concentration of the first antigenic solute to the second antigenic solute in the sample, a significant number of the ligand molecules become attached to the solid phase via the second antibody yet are capable of binding to a labelled antibody having the same specificity as the first antibody. The presence of labelled antibody bound to the solid phase support is indicative of the above mentioned situation. A method is described using the assay for determining the fertile period of the menstrual cycle by monitoring the relative concentration of oestrone-3-glucuronide and preganediol-3-glucuronide in urine. A reagent kit for use in performing the assay and method is described. A ligand molecule is described which comprises two, different antigenic radicals bound together through a bridging support molecule in which the bridging support molecule is a divalent radical derived from an organic compound having two reactive functional groups.
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