The method is useful for quantitative evaluation of bead-based lab-on-a-chip affinity assay fluorescence measurements. Fluorescent beads coated with probe molecules are immobilized (11) on a substrate. The fluorescence of the immobilized beads is measured (12) and represents the number of beads. The beads are then perfused with an analyte solution such that analyte molecules are bound to the beads. The analyte molecules are labeled with a second, different fluorescent marker which is used in a second fluorescence measurement (14) to determine the number of analyte molecules bound to the beads. A calculation (15), e.g., a quotient formation, is performed with the first (12) and the second (14) fluorescence measurement.