Using Vascular Landmarks to Orient 3D Optical Coherence Tomography Images of the Mouse Eye

Mark P. Krebs1

1 The Jackson Laboratory, Bar Harbor
Publication Name:  Current Protocols in Mouse Biology
Unit Number:   
DOI:  10.1002/cpmo.32
Online Posting Date:  September, 2017
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Comparing 3D structural information obtained by optical coherence tomography (OCT) requires accurate alignment of images acquired from individual subjects. Despite the widespread use of OCT to image the anterior and posterior mouse eye, few approaches to align the resulting image data have been described, in part due to a lack of well‐characterized landmarks that are suitable for alignment. Here, we provide an OCT acquisition and analysis protocol that incorporates the use of the long posterior ciliary arteries as landmarks. In mammals, these two large choroidal vessels lie in a plane approximately parallel to the horizon. Our OCT imaging approach resolves these vessels in the mouse eye and suggests that their location is reproducible. The protocol may be useful for preparing 3D OCT data to compare experimental cohorts of mice and for standardizing results from independent research laboratories. © 2017 by John Wiley & Sons, Inc.

Keywords: mouse models; noninvasive imaging; optical coherence tomography; Fiji/ImageJ analysis; atlas landmarks

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Table of Contents

  • Introduction
  • Strategic Planning
  • Basic Protocol 1: Three‐Dimensional Optical Coherence Tomography of the Posterior Mouse Eye
  • Support Protocol 1: Measuring Retinal Thicknesses Following Rotational Correction
  • Support Protocol 2: Adjusting the Acquisition Angle to Align LPCAs Horizontally
  • Support Protocol 3: Rotating 3D OCT Data for Analysis in ImageJ/Fiji
  • Commentary
  • Literature Cited
  • Figures
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Basic Protocol 1: Three‐Dimensional Optical Coherence Tomography of the Posterior Mouse Eye

  • Mice to be studied
  • 1% atropine sulfate ophthalmic solution drops (Akorn)
  • Ketamine/xylazine cocktail for anesthesia [1.6 ml ketamine (100 mg/ml), 1.6 ml xylazine (20 mg/ml), and 6.8 ml sodium chloride (0.9% w/v)]
  • GenTeal Severe Dry Eye Relief (Alcon Laboratories)
  • Systane Ultra Lubricant Eye Drops (Alcon Laboratories, Fort Worth, TX)
  • Bioptigen Envisu R2200 spectral domain OCT imaging system (Leica Microsystems GmbH)
  • InVivoVue Software
  • Personal computer (Mac OS X or Windows‐based) with good graphics capability, 64‐bit
  • Installed Fiji freeware, accepting all updates except developer tools on installation
  • Gloves for handling mice
  • Animal scale
  • 1‐ml syringes and 26‐G needles for injecting anesthesia
  • Sugi surgical spears (Kettenbach LP)
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Literature Cited

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  Collin, G. B., Hubmacher, D., Charette, J. R., Hicks, W. L., Stone, L., Yu, M., … Nishina, P. M. (2015). Disruption of murine Adamtsl4 results in zonular fiber detachment from the lens and in retinal pigment epithelium dedifferentiation. Human Molecular Genetics, 24(24), 6958–6974. doi: 10.1093/hmg/ddv399.
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Internet Resources
  This site may be used to install the image analysis software Fiji.
  This site provides a manual describing how to program macros in ImageJ/Fiji.
  This site may be used to install the OCT Volume Averager plugin from the Fiji update site. Updates may be added to any Fiji installation by launching the ImageJ updater with Help>Update Fiji, choosing Manage Update Sites, and selecting the JAX Eye update. Following the update, OCT Volume Averager will appear on the Plugin dropdown window of Fiji.
  This site provides the source code corresponding to OCT Volume Averager available at GitHub. It is unnecessary to obtain this file unless further development of the plugin is desired.
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