This work presents a new perspective for fluorescence signal detection using specific optics on Lab-on-Chip devices. An apparatus was designed and implemented in order to assess the performance of a fluorescence technique using different detection spot configurations, using the chip itself as a waveguide for illumination. The experiments conducted investigate the influence of the dimensions - diameter and height - of the spot on the amplitude of the detection output signal. Results show that the configuration of optical interfaces must be considered in order to improve detection output, or to be able to detect less fluorophore molecules in the spot.