Multiphoton upgrade of confocal microscope for live organ and tissue imaging

点击翻译按钮获取中文摘要

Two-photon microscopy allows for the visualization of cellular processes, such as cell migration, cell-to-cell and cell-to-matrix interactions, subcellular organelle function (mitochondria), cellular signaling (Ca2+, reactive oxygen species, and phospholipid uptake), characterization of tumor growth, drug targeting, and protein expression within the three-dimensional, natural environment of intact organs in living animals. During the past few years, two-photon microscopy has revolutionized our view of developmental, homeostatic, and pathologic processes. We propose to establish an intravital two-photon microscopy system at The Institute for Environmental Medicine (IFEM) through the acquisition of a Leica TCS SP5 Spectral Confocal and Two-Photon Microscope in combination with a Spectra-Physics Broadband Mai-Tai femtosecond-pulsed Laser. The major users of the proposed instrument will be a group of School of Medicine Investigators drawn from a wide range of disciplines, including Physiology, Pathology, Emergency Medicine, Pulmonary Medicine, Cancer Biology, Rheumatology, Hematology/Oncology and Immunology research. The need to translate experimental data from culture models into living animals, and to expand static information of dynamic cellular processes into four- dimensional kinetic analyses in vivo has greatly augmented the need for intra-vital imaging at the IFEM. Such research cannot be conducted by equipment currently available. Promising preliminary studies documenting ROS production in the lung, vascular targeting of nanocarriers, trafficking of T-cell progenitors in vivo, and immune cell interactions during inflammatory and infectious diseases has demonstrated the feasibility of two-photon microscopy for the proposed projects. The instrument will be centrally located in the School of Medicine to enable convenient access for all users, will be maintained by the IFEM under the supervision of the PI and Microscope Advisory Committee. Investigators from diverse fields will have new opportunities for interaction and the initiation of interdisciplinary research projects, which will significantly advance their NIH-funded projects. It is anticipated that the instrument will help to obtain novel insights into the normal cellular physiology, drug delivery, and various pathologies involved with aging, cancer, acute lung injury, transplantation, pulmonary hypertension, atherosclerosis, autoimmunity, and chronic infections. Relevance: The purpose of the 2-Photon microscope is to facilitate increased interaction among clinical investigators and basic scientists who are dedicated towards the study of various disease models and their possible treatment. This instrument will be invaluable in translating in vitro cell culture studies to intact organ or live animal. Overall, the equipment will enhance and expand IFEM Core to provide state of the art support to the School of Medicine at Penn.