Developmental staging is indispensable to study developmental pathways, genetics and evolution among taxa. Life-history stages have been previously described for few Poeciliid, however, proposed classifications are not generally applicable due to divergent morphology among different Poeciliid species. In Poeciliid, with placentation developmental strategy, embryos receive the maternal investment throughout gestation.Therefore, normal morpho-physiological characteristics are often compromised by applying previously employed in vitro culture and embryo fixation approaches. Here, we describe the early development of the embryonic process in a livebearing fish, Gambusia holbrooki, based on detailed morphological diagnostics and overlay cardiac rate in embryos (post heart development) under normal conditions. Light microscopy observations of live embryos obtained from gravid wild captured females, allowed assignment of the development process into seven periods: zygote, cleavage, blastula, gastrula, segmentation, pharyngula, and hatching. Thirty-seven stages were determined and assigned within each of these periods applying a numbered staging system. At least one morphologic landmark for each of the embryonic stages was defined to maximize the accuracy of staging. The heart rates showed significant differences between stages. The novelty of the present study is the real-time, in vivo quantification of the heart rhythm variations and its application as a physiological marker for embryonic staging.