Bioluminescence Imaging in an EAE Mouse Model

Abstract

Source: Schmitz, K., et al., Bioluminescence and Near-infrared Imaging of Optic Neuritis and Brain Inflammation in the EAE Model of Multiple Sclerosis in Mice. J. Vis. Exp. (2017).

This video demonstrates the method for visualizing and monitoring Experimental Autoimmune Encephalomyelitis or EAE-induced inflammation in the optic nerve using bioluminescence imaging. It includes positioning the mouse, reagent injection, and capturing sequential images to monitor changes in bioluminescence over time.

Protocol

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.

1. Bioluminescent Imaging of Optic Neuritis and Brain Inflammation

1. Setup of the imaging system
   1. Perform in vivo imaging with any equipment that allows for the analysis of bioluminescence and near-infrared signals.
   2. Keep mice under 2 - 2.5% isoflurane anesthesia during all imaging procedures.
   3. Position one or two mice beside each other in the apparatus using the middle gas supplies. Position the upper spine in the center.
   4. To evaluate medication effects, compare pairs of mice using two mice simultaneously, one per group. This is important for bioluminescent imaging.
   5. Shield the site of immunization with black cloth and take a photograph and baseline image to assess the correct positioning of the mouse/mice. For all images, use the B-focus at a 6.5 cm distance from the camera.
2. Injection and imaging of bioluminescent inflammation probe
   1. Use the in vivo imaging system settings: Epi-BLI, Em filter open, Ex filter block, fstop 1, binning 8, focus B = 6.5 cm, ad exposure 120 s; take a baseline image.
   2. Inject 100 µL i.p. of the ready-to-use chemiluminescent reagent (40 mg/mL). Mix well before filling the syringe.
   3. Capture bioluminescence images 5, 10, and 15 min after injection. The time course of the bioluminescent peak differs between animals.
      NOTE: The peak will occur 5 - 10 min after injection; a decline at 15 min indicates that no further images are required. Use mouse pairs of control and treatment groups to eliminate minor biases due to different time courses.
   4. Fill in descriptions relevant to the experiment. Observe a dialog box pop up automatically; it includes information, such as mouse strain, sex, time point, time of probe injection, group, etc. Save the files all in one folder; they will have time tags and all descriptions.

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
XenoLight RediJect Inflammation Probe	Perkin Elmer, Inc., Waltham, USA	760535	Imaging probe, activatable by oxidases, useful for imaging of inflammation
PLP139-151/CFA emulsion	Hooke Labs, St Lawrence, MA	EK-0123	EAE induction kit
IVIS Lumina Spectrum	Perkin Elmer, Inc., Waltham, USA		Bioluminescence and Infrared Imaging System
LivingImage 4.5 software	Perkin Elmer, Inc., Waltham, USA	CLS136334	IVIS analysis software
Isoflurane	Abbott Labs, Illinois, USA	26675-46-7	Anaesthetic