Colour discrimination in humans is accomplished by comparing the information from three different photoreceptor classes with distinct spectral sensitivities. Alternation or loss of one or more photoreceptor classes results in colour vision deficiency or colour blindness that affects a significant proportion of population. Its diagnosis is based on rather subjective psychophysical tests. Electroretinography (ERG) is a noninvasive and objective method for the quantitative description of cone system functionality, but no standardisation for clinical use has been made. Thus, we developed a light source, based on the synthesis of monochromatic light emitting diodes by means of a diffraction grating (LED light synthesiser), which enables spectral sensitivity measurements in less than one second with fast and reliable averaging of a large number of responses. Spectral sensitivity curves were obtained in six healthy subjects and two with colour vision deficiencies. The sensitivity curves of all healthy subjects tightly matched the photopic luminosity function standards. In subjects with colour vision deficiency, a clear deviation of curve was observed, consistent with the type of colour vision defect. Protan-type subject showed a decrease in sensitivity above 550 nm and deutan-type subject between 470 and 540 nm. The ERG spectral sensitivity measurement with the newly developed LED light synthesizer allows a fast and objective definition of human cone sensitivity and is expected to be a sensitive test for the evaluation of colour vision deficiencies.