Since the derivation of the first human embryonic stem cell (hESC) lines by Thomson and coworkers in 1998, more than 1,200 different hESC lines have been established worldwide. Nevertheless, there is still a recognized interest in the establishment of new lines of hESC, particularly from HLA types and ethnic groups currently underrepresented among the available lines. The methodology of hESC derivation has evolved significantly since 1998, when human LIF (hLIF) was used for maintenance of pluripotency. However, there are a number of different strategies for the several steps involved in establishing a new line of hESC. Here we make a survey of the most relevant parameters used between 1998 and 2010 for the derivation of the 375 hESC lines deposited in two international stem cell registries, and able to form teratomas in immunocompromised mice. Although we identify some trends in the methodology for establishing hESC lines, our data reveal a much greater heterogeneity of strategies than what is used for derivation of murine ESC lines, indicating that optimum conditions have not been consolidated yet, and thus, hESC establishment is still an evolving field of research.