Mouse cardiac fibroblasts (FBs) have been widely used as in vitro models for studying fundamental biological processes and mechanisms underlying cardiac pathologies, as well as identifying potential therapeutic targets. Cardiac FBs preparations are relatively easy to culture in a dish and can be manipulated using molecular and pharmacological tools. Because FBs rapidly decrease cell cycle division and proliferative rate after birth, they are prone to undergo phenotypic changes and senescence in cell culture as soon as a few passages. Therefore, primary cultures of differentiated FBs with amplified cell cycle from embryonic animals are required for accurate and reliable experiments. Below, we will describe a method that provides good cell yield, quality, and viability of mouse embryonic cardiac fibroblasts (MEFs) primary cultures from E16 CD-1 mice.