Thermal waters in the Yellowstone Park, WY, USA, ecosystem have created a diverse array of extreme microbial habitats. Microbial ecology studies have begun to show the diversity of extremophiles present. Focusing attention on use of a variety of collection sites, sampled materials, approaches to preservation immediately after collection, pretreatments and, perhaps most importantly, media components, will ensure a greater range of organisms in a culture collection. The objective here is to review methods for cultivating extremophiles from Yellowstone geothermal habitats.