ABSTRACT: TR&D-1 To enable beamtime at the oversubscribed LCLS source to be used for scientific discovery, not experiment optimization, the activities under this TR&D are aimed at developing the tools necessary to fully prepare for biomedical experiments and to characterize samples both before and during LCLS beam time. This ranges from understanding the sample quality and properties, as well as the state of the sample during complex sample delivery, or during a dynamic process being studied. The existing capabilities of SLAC will be enhanced via this TR&D, providing new tools in sample preparation and characterization, as well as new diagnostics to understand the state of the sample. Furthermore, capabilities at SSRL will be leveraged and augmented to use the synchrotron X-ray beam as a key tool to prepare for LCLS beamtime and for new science. Finally, spectroscopic capabilities for sample characterization at LCLS will be expanded. The proposed TR&D scope is at the cutting-edge of the biomedical structural biology field by exploiting X-ray FEL capabilities to solve new structures and study the dynamics of molecules under near-physiological conditions at ambient temperatures. The proposed developments will make use of state-of-the-art X-ray FEL capabilities and complement them with offline capabilities that themselves represent cutting-edge advances. The TR&D will increase the combined technologies’ impact on biomedical research by providing higher readiness levels to LCLS experiments for higher quality data in a shorter time. By increasing efficiency, the “extra” beam time access will be used for additional higher-risk, higher-reward experiments. It will also allow more rapid access to solve emerging scientific questions. By taking X-ray FEL sample preparation and delivery from an art to a quantitative science, it is expected that LCLS and this BTRR will have a significant impact in the structure determination of complexes and membrane proteins, provide accurate active site structures of metalloenzymes, and provide enhanced and more broadly available capabilities to observe macromolecular dynamics using the combined powers of LCLS and SSRL.