Transient expression provides a rapid, flexible and economical protein production alternative to the time consuming and costly process of generating and selecting stable high-expressing cell lines. However, there is a growing demand for higher protein yields necessary to feed the increasing needs of pharmaceutical research. This study shows how redesigning the transient expression model enabled protein yields and reproducibility, more typical of stable cell lines, while maintaining the simplicity and ease of use associated with current transient expression systems. The combination of a unique high-capacity medium, a new 293F cell variant, and the ExpiFectamine293™ transfection reagent and enhancer, together enable the new Expi293™ system to achieve expression levels approaching and exceeding 1 gram per liter of protein within 7 days. We present data demonstrating the ability of the new medium to support the increased growth rate, transfection and higher levels of protein productivity of the new Expi293F™ cell line. In some cases, expression of both IgG and non-IgG proteins were increased up to 12-fold over levels obtained with the FreeStyle™ 293 expression system. Proteins produced in the Expi293™ system exhibit biologically relevant target recognition, binding and native pharmacokinetic performance. The system is scaleable, and as a result of the high yields, enables the use of microplates for high throughput applications. The system has been demonstrated to maintain high yields with a wide variety of proteins and applications including lentiviral and membrane protein production. By enabling the generation of substantially higher levels of protein in smaller culture volumes, the Expi293™ system represents a powerful, economical and unique expression technology for both research and production of therapeutic candidate proteins.