Probing events with single molecule sensitivity in zebrafish and Drosophila embryos by fluorescence correlation spectroscopy.
Zebrafish and Drosophila are animal models widely used in developmental biology. High-resolution microscopy and live imaging techniques have allowed the investigation of biological processes down to the cellular level in these models. Here, using fluorescence correlation spectroscopy (FCS), we show that even processes on a molecular level can be studied in these embryos. The two animal models provide different advantages and challenges. We first characterize their autofluorescence pattern and determine usable penetration depth for FCS especially in the case of zebrafish, where tissue thickness is an issue. Next, the applicability of FCS to study molecular processes is shown by the determination of blood flow velocities with high spatial resolution and the determination of diffusion coefficients of cytosolic and membrane-bound enhanced green fluorescent protein-labeled proteins in different cell types. This work provides an approach to study molecular processes in vivo and opens up the possibility to relate these molecular processes to developmental biology questions.
Shi
Xianke
X
Department of Chemistry, National University of Singapore, Singapore.
Teo
Lin Shin
LS
Pan
Xiaotao
X
Chong
Shang-Wei
SW
Kraut
Rachel
R
Korzh
Vladimir
V
Wohland
Thorsten
T
D000818:Animals; D030801:Animals, Genetically Modified; D004058:Diffusion; D004331:Drosophila melanogaster / Q000196:embryology* / Q000235:genetics / Q000378:metabolism; D004625:Embryo, Nonmammalian / Q000098:blood supply; D047108:Embryonic Development / Q000502:physiology*; D005260:Female; D005453:Fluorescence; D005455:Fluorescent Antibody Technique / Q000295:instrumentation / Q000379:methods; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D008297:Male; D008954:Models, Biological; D012039:Regional Blood Flow / Q000502:physiology; D012680:Sensitivity and Specificity; D013050:Spectrometry, Fluorescence / Q000379:methods; D015027:Zebrafish / Q000196:embryology* / Q000235:genetics / Q000378:metabolism
C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins