A diverse user-group of 5 investigators, comprising The Johns Hopkins University Schools of Arts & Sciences, Engineering, and the School of Medicine, request funds to purchase a PicoQuant LSM FLIM upgrade for a Carl Zeiss LSM 780 confocal microscope. This system integrates a 1) PicoQuant LSM FLIM upgrade; 2) a 485nm pulsed diode laser for FLIM and time-correlated single photon counting; and 3) SymPho Time-64 software. The resulting state-of-the-art workstation will be sited in the Hopkins' Integrated Imaging Center (IIC, www.jhu.edu/iic) on the Homewood campus; and will represent a significant upgrade to the Hopkins-wide live imaging capabilities allowing for real-time measurements of molecular complexes within cells. The PicoQuant system will be particularly useful studying trafficking and signaling processes within cells; and will dramatically improve the sensitivity/quantification of FRET, FLIM and FCS experiments. The IIC is a Homewood campus/Hopkins-wide microscopy resource, jointly supported by the Krieger School of Arts and Sciences and the Whiting School of Engineering (see Ball & Douglas letter, page 2 of the appendix); and utilized regularly by multiple schools and departments comprising >150 laboratories and >450 users. Our investigators, all well-funded through the NIH, work on a host of disparate biological questions that include but are not limited to: RNA-protein complex assembly; lipid bilayer mediation of protein-protein interactions; trafficking of GPI proteins; PIP regulation; gene expression; biophysical mechanisms of epigenetics; nano- biosensors; molecular mechanisms of endocytosis; gene regulation; and bacterial cytokinesis. In the requested configuration, the PicoQuant system will afford our investigators the capability to precisely measure a host of molecular interactions/cellular processes. The new system will be incorporated into the IIC's existing, well established recharge system to ensure recovery of funds for supplies and maintenance; it will be made freely available to all interested users Hopkins- wide through our convenient web-scheduler; and it will be incorporated into the IIC's annual undergraduate/graduate course offerings. (www.jhu.edu/iic/academic.htm).
The PicoQuant LSM FLIM upgrade integrated with an LSM 780 confocal will be used to investigate a host of basic biomedical research questions including, but not limited to: RNA-protein complex assembly; lipid bilayer mediation of protein-protein interactions; trafficking of GPI proteins; PIP2 regulation; gene expression; biophysical mechanisms of epigenetics; nano-biosensors; molecular mechanisms of endocytosis; gene regulation; and bacterial cytokinesis. In its resulting configuration, the LSM 780/PicoQuant system will afford our investigators the capability to quantifiably measure a host of molecular interactions/cellular processes using methodologies not currently available at the Johns Hopkins University.
