Normalization of signal against reference
I assume your reference image data y_reference is acquired when there is no light on the image sensor - that's the standard thing to do. Then the usual normalized signal is
y_normalized = (y_signal - y_reference) / y_max
where y_signal is the signal image data and y_max is the maximum possible pixel output. y_max can be measured by increasing the light level on the image sensor until the output no longer increases.