Prospects for the phased anaerobic treatment of wastewater are extremely promising. With the variety of reactor designs available and the amenability of reactors to modification, existing treatment systems may be replaced or upgraded as required to achieve increased stability, higher loading capacities and greater process efficiencies than are possible using single-stage systems. In recent time, various reactor configurations and substrates are being applied to two-phase anaerobic process. This paper reviews applications and investigations of two-phase anaerobic digestion for wastewater treatment, sums up the performance of application to treating waste from distillery, landfill leachate, coffee, cheese whey and dairy, food, pulp and paper, sludge and solid etc., and summarizes reactor configurations, environmental and operational conditions, and comparisons of two-phase anaerobic digestion with other anaerobic reactors. Phase separation of anaerobic process has following major advantages: isolate and optimize potential rate-limiting steps (hydrolysis encouraged during first-phase, methanogenesis encouraged during second-phase); improve reaction kinetics and stability (pH control in each phase, improved reactor stability to shock loads, selection for faster-growing microbes); potential for detoxification in first-phase. In the other hand, phase separation has following disadvantages: disruption of syntrophic relationships; more difficulty to implement, engineering and operation; lack of process experience and applicability to variety of wastes; uncertainty of linkage between substrate type and reactor configuration. In general, substrate type and reactor configuration would determine whether or not the wastewater was amenable to two-phase digestion. There are many applications and investigations of two-phase anaerobic digestion. However, there are not enough process experience and applicability to variety of wastes. There will be much work to do.