Fluorescent Marker Reader with Uniform Illumination Calibration

The patent replaces the mechanical scanning probe system with a static wide-field illumination and imaging system. Instead of moving a single-point probe across the substrate, the invention uses a broad excitation source to illuminate the entire test region simultaneously and a sensor array to capture fluorescent emission from all regions at once. This eliminates mechanical movement while maintaining measurement capability, thereby reducing measurement time without sacrificing precision through the use of calibration-based correction algorithms.

The patent applies preliminary calibration action by first characterizing the spatial non-uniformity of the excitation illumination and detector sensitivity across the field of view. This calibration data, stored as correction factors or lookup tables, is then applied during actual measurements to compensate for position-dependent variations. This preliminary characterization eliminates measurement errors caused by non-uniform fluorophore distribution and detector response variations, significantly improving both reliability and precision.

The patent implements feedback through a calibration process where the system first measures the actual illumination profile and detector response across the field of view using a uniform fluorescent reference. These measurements feed back into the data processing algorithm, which applies position-dependent correction factors to subsequent measurements. This feedback loop enables the static wide-field system to achieve uniform measurement accuracy across the entire test substrate despite inherent non-uniformities in illumination and detection, thereby maintaining high precision while achieving fast single-shot measurements.