Visualizing morphogenesis in transgenic zebrafish embryos using BODIPY TR methyl ester dye as a vital counterstain for GFP.
Green fluorescent protein (GFP) technology is rapidly advancing the study of morphogenesis, by allowing researchers to specifically focus on a subset of labeled cells within the living embryo. However, when imaging GFP-labeled cells using confocal microscopy, it is often essential to simultaneously visualize all of the cells in the embryo using dual-channel fluorescence to provide an embryological context for the cells expressing GFP. Although various counterstains are available, part of their fluorescence overlaps with the GFP emission spectra, making it difficult to clearly identify the cells expressing GFP. In this study, we report that a new fluorophore, BODIPY TR methyl ester dye, serves as a versatile vital counterstain for visualizing the cellular dynamics of morphogenesis within living GFP transgenic zebrafish embryos. The fluorescence of this photostable synthetic dye is spectrally separate from GFP fluorescence, allowing dual-channel, three-dimensional (3D) and four-dimensional (4D) confocal image data sets of living specimens to be easily acquired. These image data sets can be rendered subsequently into uniquely informative 3D and 4D visualizations using computer-assisted visualization software. We discuss a variety of immediate and potential applications of BODIPY TR methyl ester dye as a vital visualization counterstain for GFP in transgenic zebrafish embryos.
Cooper
Mark S
MS
Department of Biology and Center for Developmental Biology, Box 351800, University of Washington, Seattle, WA 98195, USA. mscooper@u.washington.edu
Szeto
Daniel P
DP
Sommers-Herivel
Greg
G
Topczewski
Jacek
J
Solnica-Krezel
Lila
L
Kang
Hee-Chol
HC
Johnson
Iain
I
Kimelman
David
D
D000818:Animals; D030801:Animals, Genetically Modified; D001896:Boron Compounds / Q000494:pharmacology*; D004396:Coloring Agents / Q000494:pharmacology; D004952:Esters / Q000494:pharmacology; D005821:Genetic Techniques; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000494:pharmacology; D008164:Luminescent Proteins / Q000737:chemistry; D018613:Microscopy, Confocal / Q000379:methods*; D008856:Microscopy, Fluorescence; D012984:Software; D013053:Spectrophotometry; D013997:Time Factors; D019076:Transgenes; D015027:Zebrafish
C095489:4,4-difluoro-4-bora-3a,4a-diaza-s-indacene; D001896:Boron Compounds; D004396:Coloring Agents; D004952:Esters; D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins