There is a growing international scientific interest in the oil sands tailings ponds (OSTP) in northeastern Alberta, Canada, because of their significant environmental footprints, enormous volumes, and large surface areas on the affected landscape. Microbes indigenous to OSTP have several detrimental and beneficial effects in situ: they generate greenhouse gases in subsurface layers and conversely may oxidize methane at the surface; they alter pore water quality and modify tailings behavior (e.g., accelerate consolidation) through biogeochemical transformation of minerals; and they potentially mitigate OSTP toxicity through aerobic and anaerobic biodegradation. Thus, fundamental microbial processes are being investigated under different redox conditions to develop strategies for effective management of OSTP. However, the study of OSTP samples is difficult for several reasons including restricted access for sampling, spatial heterogeneity in the ponds, low biomass, and abundance of chemicals and minerals that impede microscopy and molecular biology analyses. This chapter highlights issues associated with sampling of OSTP and describes cultivation approaches and culture-independent molecular biological methods that have been used in the laboratory to study microbes and their activities in such samples, with emphasis on the sediment component of semifluid oil sands tailings.