An electrophoretic system having a plurality of separation lanes is provided with an automatic color calibration feature in which each lane is separately calibrated. For each lane, a dye matrix standard is created using reference fragments which migrate either before, or after, the sample fragments being electrophoresced, during the same electrophoresis run. These dye standards are detected automatically for the purpose of determining coefficients used to identify sample fragments mixed together with the reference fragments. This allows for calibrating the dye standard matrix for each of a plurality of lanes each time the electrophoresis system is used.