Jeffrey Perkel has been a scientific writer and editor since 2000. He holds a PhD in Cell and Molecular Biology from the University of Pennsylvania, and did postdoctoral work at the University of Pennsylvania and at Harvard Medical School.
The route a drug takes from bench to bedside can be long and arduous, and it typically passes through animals—often, a lot of animals. That’s because, in the traditional workflow, each animal represents a single dose and a single time point. With multiple doses being tested and multiple time points per dose—and of course a matching placebo group—dozens of animals might be required for even the smallest studies.
Today, researchers have a simpler, more budget-friendly option: whole-body, noninvasive preclinical imaging. Rather than sacrificing animals at each time point, and dealing with the inevitable inter-animal variability, whole-body imaging allows researchers to use fewer animals and study them repeatedly over time to produce a “longitudinal” dataset, explains Bob Pedersen, preclinical imaging product specialist at Spectral Instruments Imaging Inc. “You get very tight data.”
Preclinical tools
Preclinical imaging encompasses a range of modalities, including both anatomic (X-ray, computed tomography (CT) and magnetic resonance imaging (MRI), for instance) and molecular modalities (such as positron emission tomography (PET) and optical imaging). But as Alexandra De Lille, director of technical applications for in vivo imaging at Caliper Life Sciences, a Revvity Company, explains, optical imaging—in which fluorescent or bioluminescent probes are detected through an animal’s skin—is proving increasingly popular, being safe (as it requires no radioisotopes), easy to learn and relatively inexpensive.
Some 2,000 labs worldwide have installed a Revvity (Xenogen) IVIS optical imager, De Lille says. This has in turn generated more than 7,000 publications, mostly in the fields of oncology, immunology and neuroscience. (Revvity acquired Caliper, which previously had acquired Xenogen, in 2011.) “That clearly illustrates the acceptance of that technology in standard laboratory practice,” she says.
A number of companies have commercialized preclinical optical imagers. Besides Revvity and Spectral Instruments Imaging, there are Berthold Technologies, Bruker, LI-COR Biosciences, TriFoil Imaging and UVP. (Another company, FUJIFILM VisualSonics, offers a related technology, photoacoustic tomography, in which photo excitation of a molecular target is detected by the ultrasonic signature that event produces.)
Although each vendor has its own unique design, preclinical optical imagers tend to conform to the same basic mold: a sensitive camera mounted inside a light-tight box outfitted with one or more excitation sources. Typically, these systems also incorporate features for animal comfort, such as a heated surface and a way to administer and control anesthesia. At that point, however, the systems diverge.
Revvity’s IVIS line, for instance, includes both the IVIS Spectrum and the IVIS Lumina. The IVIS Lumina produces two-dimensional optical and X-ray images (the latter for anatomic registration), flattening the resulting positional information into a planar image, like a photograph. As a result, De Lille explains, it can be difficult to map precisely from where in the body a signal originates as well as its intensity: “Superficial” source signals tend to be overestimated, and deep-body sources are underestimated. A sufficiently deep source may be overlooked altogether.
The Spectrum systems, in contrast, produce three-dimensional datasets. Leveraging technology from two other Revvity acquisitions, VisEn and CRI, the Spectrum integrates three-dimensional X-ray (CT) data with 3D optical information that’s produced, De Lille says, by recording the relative intensity of different wavelengths of light at each pixel.
Illuminating the way
“Tissue becomes much more transparent above 600 nm,” De Lille explains. Thus, light toward the red end of the spectrum tends to pass through tissue, while bluer wavelengths are scattered and absorbed. (You can see this in action by shining a flashlight through your hand; although the beam is white light, what emerges is mostly red.) By scanning an animal over a range of wavelengths and recording the resulting spectra, the IVIS Spectrum distinguishes deep and superficial light sources essentially using the ratio of red and green signals. The resulting depth measurements are accurate to plus or minus 1 mm, De Lille says.
A more recent addition to Revvity’s optical imaging line-up is Solaris, an “open-air fluorescence imaging system” intended for fluorescence imaging in small-animal surgical applications, among others.
Spectral Instruments Imaging distinguishes its optical imagers, the benchtop Ami and floor-standing Lago, by their expanded field of view and variety of excitation sources, says Pedersen.
With a chamber measuring 25 cm on a side and 17 cm deep, the Ami “enables not just higher throughput [in terms of numbers of animals that can be imaged simultaneously], but also provides flexibility in the kinds of animals that can be imaged,” Pedersen says. Customers have used the system to image rats, vegetables and even marmosets in the Ami, he says. The Lago is even larger, with a 25-cm-deep chamber spacious enough for multiple rats.
Instead of a single white light or laser for excitation, the Ami and Lago use multiple bright LEDs ranging from blue to near-infrared: 10 LEDs and 10 emission filters on the Ami and 14 LEDs and 20 filters on the Lago. (An optional X-ray capability is also available on both systems.) The advantage, Pedersen explains, is that white-light sources, with their broad spectral excitation properties, can yield “light pollution” in the excitation spectrum even when using excitation filters. “By having a narrow wavelength of excitation light, you don’t have to throw away the polluting light; you provide only the light that is relevant to the experiment.”
Take a deeper look
For those interested in a deeper look at the animals under study, UVP’s Explorer™2 offers researchers the unique ability to zoom from the macro to the micro scale, says marketing product manager Mike Capps. Researchers can use the systems to, for instance, monitor the vasculature beneath a skin flap for the transit of fluorescently labeled cells in the animal’s circulation—something not normally visible in whole-body imagers. “You can image down to a single cell and up to a whole animal and everything in between,” Capps says.
The iBox Explorer2 (and UVP’s non-micro-capable iBox Scientia™) are fluorescence-only, but a bioluminescence-optimized version of the Scientia is anticipated this year, Capps says.
LI-COR Biosciences launched its newest imager, the Pearl® Trilogy, in March 2015. Featuring a near-infrared laser excitation source and a uniform excitation setup called FieldBrite™ Xi2, the Pearl Trilogy is capable of capturing both fluorescent and bioluminescent images over a wide dynamic range without adjusting camera settings, says Jeff Harford, senior product marketing manager. (The company’s earlier Pearl Impulse was fluorescence-only, Harford notes.) As a result, experimental reproducibility is enhanced. “You never ever get saturation of the signal, so you can use the same camera settings for all acquisitions through all studies.”
As with all its imaging hardware, LI-COR designed the Pearl Trilogy to handle near-infrared fluorophores, such as its proprietary IRDye series. According to Harford, that decision makes particular sense for small-animal work. “When you’re doing small-animal imaging, you want to be in the near-infrared anyway, because visible light doesn’t penetrate tissue well, and you’re not able to detect targets in the visible region without cutting the animal open.”
Wider adoption
With so much competition in the whole-animal optical-imaging marketplace, only time will tell whether researchers will find the Pearl’s feature set compelling. But with applications ranging from oncology to stem cells and diabetes to immunology, optical imaging, it seems, is on the rise. The challenge now, says Pedersen, is to make that growth self-sustaining, engaging researchers who don’t normally use such systems in their research.
“This is a very translatable imaging technology,” he says. “But if it’s only in the hands of a small number of users, then there’s no advantage. You’ll never push the envelope of taking this to the next level.”
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