<em>In Vivo</em> Imaging Systems (IVIS) Detection of a Neuro-Invasive Encephalitic Virus

Abstract

Modern advancements in imaging technology encourage further development and refinement in the way viral research is accomplished. Initially proposed by Russel and Burch in Hume's 3Rs (replacement, reduction, refinement), the utilization of animal models in scientific research is under constant pressure to identify new methodologies to reduce animal usage while improving scientific accuracy and speed. A major challenge to Hume's principals however, is how to ensure the studies are statistically accurate while reducing animal disease morbidity and overall numbers. Vaccine efficacy studies currently require a large number of animals in order to be considered statistically significant and often result in high morbidity and mortality endpoints for identification of immune protection. We utilized in vivo imaging systems (IVIS) in conjunction with a firefly bioluminescent enzyme to progressively track the invasion of the central nervous system (CNS) by an encephalitic virus in a murine model. Typically, the disease progresses relatively slowly, however virus replication is rapid, especially within the CNS, and can lead to an often, lethal outcome. Following intranasal infection of the mice with TC83-Luc, an attenuated Venezuelan equine encephalitis virus strain modified to expresses a luciferase gene; we are able to visualize virus replication within the brain at least three days before the development of clinical disease symptoms. Utilizing CNS invasion as a key encephalitic disease development endpoint we are able to quickly identify therapeutic and vaccine protection against TC83-Luc infection before clinical symptoms develop. With IVIS technology we are able to demonstrate the rapid and accurate testing of drug therapeutics and vaccines while reducing animal numbers and morbidity. Video LinkThe video component of this article can be found at http://www.jove.com/video/4429/ Protocol 1. Animal Preparation 1. Animal arrival: Upon arrival to the animal biosafety level 2 (ABSL2) facilities, allow animals a minimum of 2 days to acclimate to their new environment. Following this rest period, inspect the animals to evaluate their health and overall physical appearance. 1. When utilizing fluorescent reporters it is important to place all animal subjects on an alfalfa free diet to limit the amount of GI autofluorescence and background signal.2. Shave animals: To improve the bioluminescent signal detected from the cranial region during imaging; shave the heads of all the animals. Anesthetize the animals with a mixture of oxygen gas and vaporized isoflurane. Once the animals are fully anesthetized, use electric clippers to shave their heads. Once finished with shaving, return the animals to their cages and observe for complete recovery from the effects of the anesthesia. 3. Bio Medic Data Systems (BMDS) transponder insertion (to take place following shaving of the mice while the animals are fully anesthetized):For a less invasive means to track temperature implant a preprogrammed BMDS wireless transponder subcutaneousl...

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