Method of determining site-specificity and kit therefor

A method of determining the binding site specificity of an analyte that binds to a ligand having at least two different binding sites, which method comprises:

• immobilizing the ligand to a solid support,
• providing a reference analyte which binds reversibly to the ligand at a binding site thereof and whose dissociation phase after interaction with the ligand is either substantially faster or substantially slower than that of the analyte,
• contacting a mixture of the analyte and the reference analyte with the immobilized ligand to permit association to the ligand, the concentration of the one of the analyte and the reference analyte that has the faster dissociation phase being sufficient to at least substantially inhibit binding to the ligand of the one having the slower dissociation phase, should the analyte and the reference analyte both bind to the same binding site on the ligand,
• stopping the contacting of the analyte and the reference analyte with the ligand to permit dissociation therefrom,
• determining a dissociation-related value for the mixture at at least one pre-selected time during the dissociation phase, and
• determining from the determined dissociation-related value or values for the mixture if the contribution to this value or values from the one of the analyte and the reference analyte that has the slower dissociation phase is suppressed, substantial suppression indicating that the analyte and the reference analyte bind to the same binding site, and substantial absence of suppression indicating that the analyte and the reference analyte bind to different binding sites.