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Nitroreductase/Metronidazole-Mediated Ablation and a MATLAB Platform (RpEGEN) for Studying Regeneration of the Zebrafish Retinal Pigment Epithelium
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Biologia
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JoVE Journal Biologia
Nitroreductase/Metronidazole-Mediated Ablation and a MATLAB Platform (RpEGEN) for Studying Regeneration of the Zebrafish Retinal Pigment Epithelium

Nitroreductase/Metronidazole-Mediated Ablation and a MATLAB Platform (RpEGEN) for Studying Regeneration of the Zebrafish Retinal Pigment Epithelium

DOI:
10.3791/63658-v

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13:12 min

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March 02, 2022

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, ,
1Department of Ophthalmology, Louis J. Fox Center for Vision Restoration,University of Pittsburgh School of Medicine, 2Earth Research Institute,University of California, Santa Barbara, 3Department of Developmental Biology,University of Pittsburgh School of Medicine

Capítulos

  • 00:05Introduction
  • 01:00Screening Zebrafish Larvae for rpe65a:nfsB-eGFP and Genetic Ablation of the Retinal Pigment Epithelium
  • 03:02Incorporating Pharmacological Treatment into Zebrafish Retinal Pigment Epithelium Ablation Protocol
  • 06:35Confocal Microscope Z-stack Image Preprocessing in FIJI (ImageJ)
  • 08:19Quantification and Visualization of RPE Regeneration Using RpEGEN Scripts
  • 10:42Results: RPE Expression, Ablation, Regeneration Quantification
  • 12:39Conclusion

Summary

Tadução automática

Tadução automática

This protocol describes the methodology to genetically ablate the retinal pigment epithelium (RPE) using a transgenic zebrafish model. Adapting the protocol to incorporate signaling pathway modulation using pharmacological compounds is extensively detailed. A MATLAB platform for quantifying RPE regeneration based on pigmentation was developed and is presented and discussed.

Tags

Keywords: ZebrafishRetinal Pigment EpitheliumRPERegenerationNitroreductaseMetronidazoleMATLABRpEGENAblationTransgenicEGFPPTUPharmacological Treatment

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