Dual-view plane illumination microscopy for rapid and spatially isotropic imaging.
Enviado por Daniel Alfonso Colón-Ramos el
Título | Dual-view plane illumination microscopy for rapid and spatially isotropic imaging. |
Publication Type | Journal Article |
Year of Publication | 2014 |
Autores | Kumar, A, Wu, Y, Christensen, R, Chandris, P, Gandler, W, McCreedy, E, Bokinsky, A, Colón-Ramos, DA, Bao, Z, McAuliffe, M, Rondeau, G, Shroff, H |
Journal | Nat Protoc |
Volume | 9 |
Issue | 11 |
Pagination | 2555-73 |
Date Published | 2014 Nov |
ISSN | 1750-2799 |
Abstract | We describe the construction and use of a compact dual-view inverted selective plane illumination microscope (diSPIM) for time-lapse volumetric (4D) imaging of living samples at subcellular resolution. Our protocol enables a biologist with some prior microscopy experience to assemble a diSPIM from commercially available parts, to align optics and test system performance, to prepare samples, and to control hardware and data processing with our software. Unlike existing light sheet microscopy protocols, our method does not require the sample to be embedded in agarose; instead, samples are prepared conventionally on glass coverslips. Tissue culture cells and Caenorhabditis elegans embryos are used as examples in this protocol; successful implementation of the protocol results in isotropic resolution and acquisition speeds up to several volumes per s on these samples. Assembling and verifying diSPIM performance takes ∼6 d, sample preparation and data acquisition take up to 5 d and postprocessing takes 3-8 h, depending on the size of the data. |
DOI | 10.1038/nprot.2014.172 |
Alternate Journal | Nat Protoc |
PubMed ID | 25299154 |
PubMed Central ID | PMC4386612 |
Grant List | OD016474 / OD / NIH HHS / United States R24 OD016474 / OD / NIH HHS / United States U01 HD075602 / HD / NICHD NIH HHS / United States U01 HD075602 / HD / NICHD NIH HHS / United States |