Fed-batch processes are a current preference for the production of recombinant proteins in mammalian cells. The use of nutrient feeding prevents the depletion of important medium components and results in improved culture longevity and high cell and product yields. To take maximum advantage of these effects, it is important to optimize the fed-batch process for each application. In this chapter, a simple strategy for fed-batch optimization is described, consisting of the development of a feed medium based on spent media analysis and the establishment of a feeding strategy that consists of adding variable volumes of feed media at specific intervals, after off-line measurement of the concentration of a reference nutrient.