Design and Construction of a Cost-Effective Spinning Disk System for Live Imaging of Inner Ear Tissue.
Confocal imaging of fluorescent probes offers a powerful, non-invasive tool which enables data collection from vast population of cells at high spatial and temporal resolution. Spinning disk confocal microscopy parallelizes the imaging process permitting the study of dynamic events in populations of living cells on the millisecond time scale. Several spinning disk microscopy solutions are commercially available, however these are often poorly configurable and relatively expensive. This chapter describes a procedure to assemble a cost-effective homemade spinning disk system for fluorescence microscopy, which is highly flexible and easily configurable. We finally illustrate a reliable protocol to obtain high-quality Ca(2+) and voltage imaging data from cochlear preparations.
Department of Physics and Astronomy, University of Padua, via Marzolo 8, Padua, 35131, Italy.|Foundation for Advanced Biomedical Research, Venetian Institute of Molecular Medicine (VIMM), via G. Orus 2, Padua, 35129, Italy.|CNR, Institute of Cell Biology and Neurobiology, via E. Ramarini 32, Monterotondo, RM, 00015, Italy.
Federico
F
Ceriani
CNR, Institute of Neuroscience, viale G. Colombo 3, Padua, 35121, Italy.
Catalin D
CD
Ciubotaru
Department of Physics and Astronomy, University of Padua, via Marzolo 8, Padua, 35131, Italy. mario.bortolozzi@unipd.it.|Foundation for Advanced Biomedical Research, Venetian Institute of Molecular Medicine (VIMM), via G. Orus 2, Padua, 35129, Italy. mario.bortolozzi@unipd.it.|CNR, Institute of Cell Biology and Neurobiology, via E. Ramarini 32, Monterotondo, RM, 00015, Italy. mario.bortolozzi@unipd.it.
Mario
M
Bortolozzi
Department of Physics and Astronomy, University of Padua, via Marzolo 8, Padua, 35131, Italy.|Foundation for Advanced Biomedical Research, Venetian Institute of Molecular Medicine (VIMM), via G. Orus 2, Padua, 35129, Italy.|CNR, Institute of Cell Biology and Neurobiology, via E. Ramarini 32, Monterotondo, RM, 00015, Italy.
Fabio
F
Mammano
D000818:Animals; D015220:Calcium Channels / Q000378:metabolism; D007758:Ear, Inner / Q000378:metabolism / Q000648:ultrastructure*; D051379:Mice; D018613:Microscopy, Confocal / Q000295:instrumentation / Q000379:methods; D008856:Microscopy, Fluorescence / Q000295:instrumentation / Q000379:methods
D015220:Calcium Channels
Calcium action potentials; Calcium imaging; Calcium signaling; Confocal spinning disk microscopy; Connexins; Inner hair cells; Non-sensory cells; Spontaneous activity; Voltage imaging