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. 2000 Oct 24;97(22):12278-82.
doi: 10.1073/pnas.97.22.12278.

Visualizing gene expression by whole-body fluorescence imaging

M Yang  1 , E Baranov, A R Moossa, S Penman, R M Hoffman
Affiliations

Visualizing gene expression by whole-body fluorescence imaging

M Yang et al. Proc Natl Acad Sci U S A. .

Abstract

Transgene expression in intact animals now can be visualized by noninvasive techniques. However, the instruments and protocols developed so far have been formidable and expensive. We describe here a system for rapidly visualizing transgene expression in major organs of intact live mice that is simple, rapid, and eminently affordable. Green fluorescent protein (GFP) is expressed in the cells of brain, liver, pancreas, prostate, and bone, and its fluorescence is encoded in whole-body optical images. For low-magnification images, animals are illuminated atop a fluorescence light box and directly viewed with a thermoelectrically cooled color charge-coupled device camera. Higher-magnification images are made with the camera focused through an epi-fluorescence dissecting microscope. Both nude and normal mice were labeled by directly injecting 8 x 10(10) plaque-forming units/ml of adenoviral GFP in 20-100 microl PBS and 10% glycerol into either the brain, liver, pancreas, prostate, or bone marrow. Within 5-8 h after adenoviral GFP injection, the fluorescence of the expressed GFP in brain and liver became visible, and whole-body images were recorded at video rates. The GFP fluorescence continued to increase for at least 12 h and remained detectable in liver for up to 4 months. The system's rapidity of image acquisition makes it capable of real-time recording. It requires neither exogenous contrast agents, radioactive substrates, nor long processing times. The method requires only that the expressed gene or promoter be fused or operatively linked to GFP. A comparatively modest investment allows the study of the therapeutic and diagnostic potential of suitably tagged genes in relatively opaque organisms.

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Figures

Figure 1
Figure 1
External and internal images of vAd-GFP gene expression in various organs. External fluorescent whole-body images were compared with direct images of vAd-GFP gene expression in various organs. Series of external fluorescence images of vAd-GFP gene expression in brain, liver, pancreas, prostate, and tibia, (A, C, E, G, and I) compared with corresponding images of the exposed organs (B, D, F, H, and J). See Materials and Methods for vector delivery and imaging details.
Figure 2
Figure 2
External whole-body image of vAd-GFP gene expression in the brain. An external image of vAd-GFP gene expression in the brain acquired from a nude mouse in the light box 24 h after gene delivery. Clear image of transgene expression in the brain can be visualized through the scalp and skull. See Materials and Methods for imaging.
Figure 3
Figure 3
Time course of GFP gene induction in the brain of a nude mouse. [I′GFP] (see Materials and Methods) from a series of whole-body images determined that vAd-GFP expression could be first visualized by 5 h, 15 min after local introduction of vAd-GFP gene in a nude mouse. Increases in expression could be visualized in 30-min intervals.
Figure 4
Figure 4
Time-course GFP gene induction in the liver of a nude mouse. vAd-GFP expression [I′GFP] (see Materials and Methods) could be first visualized in the liver by 7 h, 45 min after introduction of vAd-GFP gene in a nude mouse. Increases in expression could be visualized in 30-min intervals.
Figure 5
Figure 5
External whole-body image of vAd-GFP gene expression in the liver. An external image of vAd-GFP gene expression acquired from a nude mouse in the light box 72 h after gene delivery. Lateral, whole-body image of transgene expression in the liver can be clearly visualized through the abdominal wall.
See this image and copyright information in PMC

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