Systems and methods for producing background-reduced fluorescence imaging signals include an illumination system that provides illumination light from an illumination source to a targeted area on the sample platform, a sensor adapted to detect light and having an array of sensing locations, and collection optics arranged and configured to project light emanating from the sample platform onto the sensor. In typical operation, light from the targeted area is projected onto a first portion of the sensor comprising a first plurality of the sensing locations and light from proximal to the targeted area on the platform is projected onto a second portion of the sensor comprising a second plurality of the sensing locations, and a second signal detected by the second portion of the sensor is subtracted from a first signal detected by the first portion of the sensor to produce a background-reduced signal, e.g., a signal with reduced background related noise.
