Geotrichum candidum is a micro-fungus widely used as a ripening starter in cheese making. In anthropogenic environments such as dairy industries, this microorganism is subjected to many environmental and technological stresses including low temperature exposure. Our aim was to study the proteomic response of G. candidum to cold stress using a comparative proteomic approach by two-dimensional Differential In Gel Electrophoresis (2D DIGE). This technique consists on the labeling of proteins by specific fluorescent dyes (CyDyes). The results, obtained with G. candidum cells subjected to cold temperature, show significant proteomic patterns differences compared with the standard conditions. Furthermore, this biochemical response seems strain specific. 2D DIGE technology combined with SameSpots™ software analysis support these results through an important statistical validity. The comparative studies in a single gel, using two different fluorescent CyDyes (Cy3 and Cy5), lead to proteins differentiation. Selected spots were treated and analyzed by mass spectrometry.