@article{b9a55da847694599ba3ecb0c246e742e,
title = "A layered mounting method for extended time-lapse confocal microscopy of whole zebrafish embryos",
abstract = "Dynamics of development can be followed by confocal time-lapse microscopy of live transgenic zebrafish embryos expressing fluorescence in specific tissues or cells. A difficulty with imaging whole embryo development is that zebrafish embryos grow substantially in length. When mounted as regularly done in 0.3-1% low melt agarose, the agarose imposes growth restriction, leading to distortions in the soft embryo body. Yet, to perform confocal time-lapse microscopy, the embryo must be immobilized. This article describes a layered mounting method for zebrafish embryos that restrict the motility of the embryos while allowing for the unrestricted growth. The mounting is performed in layers of agarose at different concentrations. To demonstrate the usability of this method, whole embryo vascular, neuronal and muscle development was imaged in transgenic fish for 55 consecutive hours. This mounting method can be used for easy, low-cost imaging of whole zebrafish embryos using inverted microscopes without requirements of molds or special equipment.",
keywords = "Confocal microscopy, Developmental Biology, Embryo, Issue 155, Mounting, Time-lapse, Transgenic, Zebrafish",
author = "Sanat Upadhyay and Leoncio Vergara and Pranjali Shah and Gustafsson, {Jan {\AA}ke} and Kakadiarias, {Ioannis A.} and Maria Bondesson",
note = "Funding Information: This work was supported by grants from the National Institute of Environmental Health Sciences of the National Institutes of Health (grant number P30ES023512 and contract number HHSN273201500010C). SU was supported by a fellowship from the Keck Computational Cancer Biology program (Gulf Coast Consortia CPRIT grant RP140113) and by Hugh Roy and Lillie Endowment Fund. J-{\AA} G was supported by the Robert A. Welch Foundation (E-0004). Funding Information: We thank Albert Pan and Arndt Sieakmann for gifts of transgenic fish. We thank Koichi Kawakami at National Institute of Genetics, the National BioResource Project from the Ministry of Education, Culture, Sports, Science and Technology of Japan for the gift of the transgenic zebrafish HGn39b. We also thank Fatima Merchant and Kathleen Gajewski for assistance on confocal microscopy, and Tracey Theriault for photographs. Funding Information: We thank Albert Pan and Arndt Sieakmann for gifts of transgenic fish. We thank Koichi Kawakami at National Institute of Genetics, the National BioResource Project from the Ministry of Education, Culture, Sports, Science and Technology of Japan for the gift of the transgenic zebrafish HGn39b. We also thank Fatima Merchant and Kathleen Gajewski for assistance on confocal microscopy, and Tracey Theriault for photographs. This work was supported by grants from the National Institute of Environmental Health Sciences of the National Institutes of Health (grant number P30ES023512 and contract number HHSN273201500010C). SU was supported by a fellowship from the Keck Computational Cancer Biology program (Gulf Coast Consortia CPRIT grant RP140113) and by Hugh Roy and Lillie Endowment Fund. J-? G was supported by the Robert A. Welch Foundation (E-0004). Publisher Copyright: {\textcopyright} 2020 Journal of Visualized Experiments.",
year = "2019",
month = jan,
doi = "10.3791/60321",
language = "English (US)",
volume = "2020",
journal = "Journal of visualized experiments : JoVE",
issn = "1940-087X",
publisher = "MYJoVE Corporation",
number = "155",
}