Featured Article
Monday June 02, 2008
By Caitlin Smith
Visualize the interior of your animal model at the moment its disease really takes hold—wouldn’t it be amazing, not to mention instructional? That’s what in vivo imaging, especially with whole animals, is attempting to do. Recent success in this area holds promise for disease researchers, drug development, and clinical medicine.
Supporting in vivo imaging is a molecular toolbox with vast and varied contents. Fluorophores, which emit a particular wavelength of light upon illumination with a different wavelength of light, as well as bioluminescent molecules that emit light upon stimulation by chemical substrates, make it possible to tag or trace almost any molecule with surprising reliability. The combination of imaging and molecular tools at the whole-animal level is also attractive for its relative non-invasiveness, low toxicity, and ease of studying groups of animals together over a period of time.
Mobile x-ray systems
The utility of x-ray imaging as a research and screening tool for in vivo labs has made portable units nearly indispensable. “Planar/2-D imaging continues to remain a complementary and core screening technique for the small animal market,” says Brad Jackson, vice president of sales and marketing at Faxitron X-Ray. “This is primarily due to its relatively low price point and high throughput, two key drivers for the research market.” One example is Faxitron’s new LX-60 system for small animal imaging (mice, rats, and larger rodents). Faxitron claims to lead their competitors on spatial resolution by virtue of their unique detectors, which allow higher resolution over the entire field of view.
Historically, contrast resolution and spatial resolution have always challenged imaging scientists. Vikram Butani, director of Kubtec, says that the real challenge is improving both of these while also reducing costs. “3-D x-ray is more and more readily available today, but it is fairly expensive. A good 3-D machine is at least three times what a good standard 2-D machine costs.” Most researchers cannot afford to have a 3-D x-ray system in their lab—and they might not really need one anyway. “It’s all very exciting, with beautiful images, but there’s a lot more there than is really needed.” While some research facilities address this need by putting a 3-D system in a core imaging facility for researchers to share, Butani maintains that often this arrangement is unsatisfactory, citing limitations involving booking time, transporting animals, and vivarium requirements. “A lot of times, the images a 2-D machine can give you are more than sufficient,” says Butani. Kubtec plans to introduce a 2-D x-ray system that is more affordable than current 2-D machines, yet simultaneously offers enhanced features that are closer to that obtained with a 3-D system. “So it’s almost like getting a limited version of a 3-D machine in your own lab,” explains Butani. “That is something that we’re working on, and we’ll have it out this year.”
Kubtec’s Xpert 80L system, introduced last year, is another example of a portable 2-D system designed to reduce costs. Scientists studying nonhuman primates typically use an x-ray room, which is relatively expensive. Large enough for many types of nonhuman primates, the Xpert 80L can be wheeled between labs or around a vivarium. “And the cost of one of these with the digital detector is probably 20% of what it would cost to put a room together,” says Butani. The Xpert 80L can also be used for smaller animals, imaging animals singly or together as a group. “When you’re doing work with mice,” says Butani, “it doesn’t make sense to use an entire x-ray room.”
Better sensitivity
CRi offers a unique approach to in vivo fluorescence imaging that takes advantage of spectral imaging. They use spectral unmixing to reduce the interference of autofluorescence from surrounding tissue, similar to the concept of multiplexing, where multiple signals can be measured simultaneously if their spectra are known (and separable). If the spectra of the desired signals and the autofluorescence are known or deducible, they can be “unmixed” from one another in the fluorescent signal from the sample. “We have been shown to be up to 300 times more sensitive (i.e., with a 300 times lower limit of detection of fluorophores) than competing systems,” says James Mansfield, CRi’s director of multispectral imaging systems. “This is what makes doing in vivo fluorescence imaging practical.” Their new Maestro 2 In Vivo Imaging System also has multiplexing capabilities for studying multiple fluorescent signals.
CRi also recently introduced DyCE (for dynamic contrast enhancement), a technique that Mansfield describes as “a completely novel means of investigating biodistributions of fluorophores and generating all-optical anatomic maps of mice.” DyCE works by collecting a series of dynamic images after injecting a bolus of near-infrared dye, which circulates through the body and causes characteristic responses in different organs. The locations of major organs are derived by CRi’s proprietary algorithms. The organs are displayed along with visible changes in them as the dye circulates, resulting in anatomical maps that can be correlated with molecular imaging data for the same animal.
Mansfield believes that the biggest hurdle facing in vivo imaging researchers is 3-D optical imaging. “Some companies claim to be able to do this, but they have come out with systems that incorporate homogeneous models of mice,” he says, “and as we all know, mice are far from homogeneous.” CRi is developing a method of 3D imaging that incorporates “non-homogeneous light scattering models (supplied by DyCE anatomic images) that we feel will finally enable researchers to get valid 3D images,” says Mansfield.
Another in vivo imaging system offering spectral unmixing of fluorescent signals is the KODAK In-Vivo Multispectral Imaging System FX from Carestream Molecular Imaging. It’s the first multispectral in vivo system to also offer x-ray imaging capabilities. According to their director of research and development, William McLaughlin, “researchers can identify tumor cells with one targeting agent at one wavelength, while assessing vasculature growth surrounding the tumor with a different targeting agent at another wavelength. These images can be combined with a high-resolution x-ray image to provide much more accurate localization of the two targets.”
For improved detection in deeper tissues, LI-COR just launched the Pearl Imager for small animal imaging. “The unique aspects of the Pearl are that it is a system designed and optimized for near-infrared detection,” says Jeff Harford, LI-COR product manager. “Near infrared dyes and detection are extremely important for small animal imaging, particularly at 800 nm wavelengths.” The new release joins their 800 nm dye, IRDye 800 CW, for near-infrared detection tools.
Targeting agents and barriers
Harford says that academic and commercial efforts are expanding the molecular targeting agents for in vivo imaging. “Since the goal for molecular imaging is to be translatable to the clinic,” he says, “work is being done on multi-modal approaches, such as labeling a targeting agent with both NIR dyes as well as radio labeling for both optical detection and detection on modalities such as Spect/Pet/CT.” But Harford notes that it is still difficult to get “reliable targeting agents that are small enough to clear the animal’s system, and yet also bind specifically to the target of interest. For example, antibodies are easily labeled and used, but will generally not clear the animal’s system as well as a small peptide.” Carestream Molecular Imaging’s Kodak X-SIGHT organic fluorescent imaging agents, which are biocompatible, nontoxic, photostable nanoparticles, are a new option. “These nanoparticles deliver not only longer circulation,” says McLaughlin, “but also enhanced photostability, allowing more time for image capture.” They are available in several fluorescent wavelengths, including in the near-infrared range.
One daunting biological roadblock to reliable targeting agents is the blood-brain barrier, says Harford. Many neuroscientists are denied the benefits of in vivo imaging because they simply can’t penetrate this wall despite the burgeoning molecular toolbox now at hand. No doubt it is just a matter of time, and one day, open access to the brain by in vivo imaging will likely spawn entire new fields of research.