Fig. 1.

Fig. 1. In vivo imaging of fluorescence anisotropy controls expressed in pancreatic β cells of 5-dpf zebrafish embryos.

(A) Top: Tol2 transposon constructs used to express two fluorescence anisotropy controls, R198P and TDimer, in zebrafish. Expression is targeted to β cells using a zebrafish insulin promoter. Bottom: The R198P and TDimer establish the upper and lower bounds, respectively, of the Apollo-NADP⁺ dynamic range. pA, polyadenylation signal. (B) Transgenic zebrafish embryo at 5 dpf, imaged from the right lateral view. White arrow indicates the pancreatic islet. (C) Schematic of microfluidic device and features used to hold zebrafish embryos during imaging. (D) Schematic of microfluidic device cross section and perfusion flow through the device. PDMS, polydimethylsiloxane. (E) In vivo pancreatic islet slice images from 5-dpf R198P (top) and TDimer (bottom) transgenic zebrafish embryos, taken at various depths (5 to 40 μm). (F) In vivo fluorescence anisotropy imaging of 5-dpf islets of R198P and TDimer transgenic zebrafish embryos, taken at 1-μm intervals. n = 25 to 27 embryos. (G) In vivo time series fluorescence anisotropy imaging of a single islet slice from 5-dpf R198P and TDimer transgenic zebrafish embryos, taken at 10-s intervals. n = 13 to 15 embryos.