OPHTHALMOLOGY/MEDICAL IMAGING: Retinal imaging advances research, disease diagnosis

Recent instrumentation progress is enabling superior imaging of the retina in both laboratory animals and humans. The effect is better research for neurological disorders, for instance, as well as disease diagnosis.

The organ we most depend on for providing input about our environment–the eye– is helpful for the diagnosis of diseases such as diabetes. Also, because the retina is part of the brain, its assessment enables identification of neurological effects–and thus the eye warrants significant research attention.

A number of technology advances are fueling progress in retinal imaging–both for research and clinical application.

In vivo research

Mice have played a key role for research into biological functions such as vision, and are especially attractive for genetic research because they have a particularly "plastic" genome. They can be genetically engineered to have various diseases that mimic human dystrophies such as diabetes and Alzheimer's disease.

Click to Enlarge

Figure 1. Phoenix Research Laboratories designed its Micron III retinal imaging microscope for easy operation by a single technician.

"A barrier to effective animal research has been the difficulty of imaging the retina in these tiny mammals," says Bert Massie of Phoenix Research Laboratories (Pleasanton, CA; www.phoenixreslabs.com), explaining that the mouse eye, at 3 mm in diameter, is a fraction of the size of the 25-mm human eye. "Attempts to use cameras designed for the human eye have produced limited results and at great expense," he notes.

Addressing the great need for retinal imaging specifically in small animal research, Phoenix Research Labs provides instrumentation that incorporates multiple imaging modalities important for eye research. These include white light (also called brightfield) imaging; angiography, in which injected fluorescein enables retinal blood flow study; and imaging of fluorescent probes such as green fluorescent protein (GFP) where the challenge is to observe a low level of fluorescent return. When Phoenix Research Laboratories developed its Micron III retinal imaging microscope specifically for laboratory rodents, a key design objective was to deliver these imaging modalities–along with a wide field of view; resolutions below 5 µm; and easy, rapid imaging (a mouse or rat can be imaged by a solo technician in less than a minute). (See Fig. 1.)

Key to achieving these design objectives is the system's CCD camera. Massie explains that a standard, single-chip CCD camera could not deliver the performance needed because it uses a mask over the pixels with an array of filters to capture a color image. Since most of the spatial information is in the green, the pattern of filters provides twice the number of green samples as red or blue, and the software interpolates the colors where measurements are not made. By contrast, a three-chip CCD uses a prism to separate the colors, each of which is then sent to one of three CCDs. This allows equal spatial resolutions for each color, for good color images and for fluorescent studies at arbitrary wavelengths. The Micron III uses Toshiba Imaging's (Irvine, CA; cameras.toshiba.com) 56 × 44 × 44 mm IK-TF7 three-chip CCD camera to provide high resolution (XGA) and a full pixel-independent readout of 30 frames per second and 1000 to 1 dynamic range. Important to live animal retinal imaging tasks, three 1/3-in. progressive scan CCDs eliminate image jitter.

Another key to improved research is the ability to perform in vivo imaging of the mice. Traditionally, researchers must breed large colonies of animals, sacrifice a segment of the colony at each stage of disease progress, and then remove the retina and flat-mount it to a microscope slide for examination. The process is complicated, and precludes longitudinal studies. The Micron III addresses this need as well.

Click to Enlarge

Figure 2. Multi-modal imaging enables generation of various depictions from a single instrument: a retinoblastoma, a juvenile eye tumor as it appears in a mouse (top left); an anterior defect in an albino rat (top right); a retinal defect in a mouse depicted by an angiogram (bottom left); and GFP expression in a rat (bottom right). (Images courtesy Phoenix Research Laboratories)

The results are providing new insights into the eye. A typical set of images demonstrates the system use in a variety of observational modalities, from bright field to fluorescein angiography to GFP (see Fig. 2). New attachments are in development for imaging the anterior segment and for performing visual function tests.

Research to clinical

Imagine Eyes (Orsay, France, www.imagine-eyes.com), developer of the compact adaptive optics retinal camera prototype described in the article, "Adaptive optics approaches clinical ophthalmology," http://bit.ly/cC4gkp), says it expects to launch its second-generation system this Fall.

Click to Enlarge

Figure 3. Detected using qualitative oximetry with Optical Imaging's Retinal Functioning Imager (RFI), perfusion deficits and abnormalities in a patient with sickle cell retinopathy appear as regions of color distinct from their surroundings. (Image courtesy Richard B. Rosen and Teerapat Jittpoonkuson, New York Eye and Ear Infirmary)

The system's first generation had a prototype interface and a 4 × 4° field of view at ±2–3 µm.  The goals of the second generation are a more ergonomic interface, a wider imaging field and better image contrast. It is geared to enable clinical and academic users with minimal training to capture and stitch together component images to form large, wide-field views. "This second generation will be a crucial step in launching a commercial camera as quickly as possible," says Mark Zacharria, director of marketing communications at Imagine Eyes.

Meanwhile, the company will provide component technologies to basic research as well as systems to academic and industrial users.

Clinical functional imaging

Speaking at the BiOS Hot Topics session during Photonics West 2010, Professor Amiram Grinvald, founder and CEO of Optical Imaging Ltd. (www.opt-imaging.com) and a faculty member of the Weizmann Institute of Science (Rehovot, Israel), described the application of his company's Retinal Function Imager (RFI), an FDA-approved hardware-and-software system providing noninvasive, ophthalmic functional imaging–in < 1 s to 10 min–to the resolution of single red blood cells moving through capillaries. 1 The system is partially based on a technique described in Grinvald's 1986 paper in Nature for functional imaging of the brain based on intrinsic signals. 2 Alternative technologies, such as PET and f-MRI, still provide only 1–10% of the resolving power of Grinvald's approach in both the temporal and the spatial domains, he says.

RFI enables direct visualization of retinal blood dynamics without the injection of contrast agents, and clearly reveals the motion of individual red blood cells and blood cell clusters, thus enabling quantitative detection of abnormal blood flow velocity in capillaries, arterioles, and venules. This opens the door to many new diagnostic possibilities–for instance, a significant velocity decrease in arteries and veins may indicate non-proliferative diabetic retinopathy. And increased blood flow velocity in the retinal arteries and veins can be an early indicator of diabetes mellitus in patients without any sign of diabetic retinopathy.

The basic RFI 3000 offers both blood flow velocity mapping and capillary perfusion mapping (CPM), which enables analysis of a series of images to reveal motion and microvasculature detail–often in greater detail than the gold standard for clinical retinal imaging: fluorescein angiography (FA).


Click to EnlargeClick to Enlarge

Figure 4. Capillary perfusion mapping (CPM; left) enables better visibility of capillaries compared with the gold standard imaging method, fluorescein angiogram (FA; right). In addition, CPM is completely noninvasive, and as such it allows patient follow-up not feasible with FA after treatment with drugs or surgery. (Image courtesy Optical Imaging Ltd.)

The higher-end RFI 3006 adds a multispectral imaging and analysis module, for instance, to provide insight into oxygen use and other functions. Based on a fast-switching filter wheel, it overcomes issues such as poor signal-to-noise ratio that have typically hampered such analyses. The module enables assessment of the oximetric state of the retina and visualization of choroidal vessels. This latter option provides ICG-like images (using the contrast agent IndoCyanine-Green) without the use of ICG.

Retinal reflectance changes in response to photic stimulation carry information about metabolic processes–which is the basis for the 3006's metabolic functional imaging capability. RFI can image under near-IR light, outside the absorption range of the photoreceptors, and thus can be used to optically monitor retinal activity in response to a well-defined visual stimulus. The metabolic state of retinal components is determined by comparing post and pre-stimulated images, reflecting changes in absorption outside the absorption range of the photoreceptors or scattering. It has been found in animal model experiments that the functional signal is very sensitive for the detection of induced glaucoma.

The system features a 12.3-mm square sensor with resolution of 1024 × 1024 pixels that capture images at 60 Hz. Its light source is a stroboscopic xenon with eight flashes at 100-Hz maximum frequency and a 10-s inter-series recharging interval. Imaging optics include a Topcon TRC-50DX fundus camera with three-position variable field angle. Software modules enable capture, sophisticated analysis, storage and retrieval, image processing, and display of related imagery as well as relevant historical and clinical information.


  1. D. Izhacky et al., Jpn. J. Ophthalmol., 53(4):345–51, July 2009.
  2. A. Grinvald et al., Nature, 1986, 324: 361–364.


More Brand Name Current Issue Articles
More Brand Name Archives Issue Articles

Get All the BioOptics World News Delivered to Your Inbox

Subscribe to BioOptics World Magazine or email newsletter today at no cost and receive the latest news and information.

 Subscribe Now
Related Articles

Fluorescence expands swallowable camera capsule's cancer detection capabilities

Fluorescent light expands the diagnostic capabilities of a swallowable camera capsule for throat and gut cancer detection.

Spectral microscopy captures metal-labeled neurons in 3D, and with unprecedented detail

A team of researchers used spectral confocal microscopy to image tissues impregnated with silver or gold.

LuxCath optical tissue characterization catheter enables real-time monitoring during cardiac ablation

A study used optical tissue characterization technology for the first time in procedures to treat arrhythmia patients.

EUV spectral imaging tool can map cell composition in 3D

A newly developed spectral imaging instrument enables observation of how cells respond to new medications at a minute level.

Fluorescence Imaging: Optical filtering basics for life sciences

Optical filters can have a dramatic effect on outcomes in life sciences. These principles demonstrate how next-generation thin film enhances excitation and emission in fluorescence bioimaging syste...

Photoacoustics/Biomedical Imaging: Photoacoustic imaging progresses toward medical diagnostics

Recent technological developments in laser and transducer hardware, contrast agents, and image reconstruction algorithms have helped to advance photoacoustic (or optoacoustic) imaging.  

Translational Research: Bench-to-bedside: Progress, pioneers, and 21st Century Cures

The NIH/SPIE Biophotonics from Bench to Bedside workshop (Sept. 24-25) featured speakers and posters presenting exciting translational research in technologies and applications.

Legislation promises biophotonics opportunities

The 21st Century Cures Act (H.R. 6) was a focal point at the NIH/SPIE Biophotonics from Bench to Bedside workshop.

Zeiss partners with Molecular Imaging Platform at McGill University Health Centre

Zeiss has entered into a partnership with the Research Institute of the McGill University Health Centre's Molecular Imaging Platform.


Neuro15 exhibitors meet exacting demands: Part 2

Increasingly, neuroscientists are working with researchers in disciplines such as chemistry and p...

Why be free?

A successful career contributed to keeping OpticalRayTracer—an optical design software program—fr...

LASER Munich 2015 is bio-bent

LASER World of Photonics 2015 included the European Conferences on Biomedical Optics among its si...

White Papers

How can I find the right digital camera for my microscopy application?

Nowadays, image processing is found in a wide range of optical microscopy applications. Examples ...



Twitter- BioOptics World

Copyright © 2007-2015. PennWell Corporation, Tulsa, OK. All Rights Reserved.PRIVACY POLICY | TERMS AND CONDITIONS