http://medicalphysicsweb.org/cws/article/research/39558

Fluorescence imaging is shaping up as a promising modality for guiding surgical resection of tumours; but the technique could also prove of benefit for deep tissue imaging. Now, a research team headed up by Brian Pogue of Dartmouth College (Hanover, NH) has reported the first in vivo demonstration of deep-tissue tomographic imaging using the endogenous fluorescence of protoporphyrin IX (J. Biomed. Opt. 14 030501).

The team - from Dartmouth College, ART of Canada, the UK's University of Birmingham and Dartmouth Medical School (Hanover, NH) - coupled a fluorescence tomography system to a microCT to allow both X-ray structural and optical functional imaging. Imaging of an animal model revealed that endogenous protoporphyrin IX (PpIX) fluorescence enabled tumour imaging from within the cranium and detected glioma tumours that were not apparent in the microCT images.

"The work we are doing in combined microCT/optical tomography is an experiment in how to usefully combine X-ray CT with optical tomography in a non-contact arrangement," explained Pogue. "This is challenge, but the system is working and was intended as a demonstration prototype that might be developed into an alpha system by ART if deemed to have useful market value. We are still in the analysis phase, but all measures to date indicate it is working well."

Sensitivity boost
Development of tomographic PpIX fluorescence imaging has previously been hampered by problems such as low signal intensity and background skin fluorescence limiting sensitivity to deeper structures. To address this, the researchers employed time-correlated single-photon counting in their tomography system, enabling imaging through deep tissue in a reasonable time frame.

The system uses 635 nm excitation light from a 1-mW pulsed diode laser, collimated to a 1-mm effective area. Five detectors - time-resolved photomultiplier tubes with single-photon-counting electronics - are arranged in an arc on the opposite side of the animal being imaged, allowing non-contact measurement of the emerging light signal (the PpIX fluorescence peak near 700 nm). Scanning is enabled by rotating the beam around the animal.

Emerging signals are split into fluorescence (95%) and transmission (5%) channel detectors, enabling the fluorescence-to-transmission signal ratio to be calculated at each location. Employing such a ratio makes the data resistant to many calibration errors, distance inaccuracies and tissue heterogeneity.

The researchers tested the fluorescence tomography system on a mouse with a glioma tumour implanted in its brain. Gliomas provide significant endogenous fluorescence from PpIX, which is enhanced by administering aminolevulinic acid (ALA) to stimulate further PpIX production.

The team first imaged the tumour with contrast MRI to assess its size and location and acquired a microCT dataset (using a GE eXplore Locus SP scanner with 94-µm pixel detector resolution). They then used fluorescence tomography system to acquire a pre-ALA-injection dataset (at 64 angles and with a total scan time of 10 min). Following ALA administration and incubation for 1 hr, the animal was re-imaged to generate a post-contrast dataset.

The fluorescence-to-transmission ratio data were reconstructed into an image, which the researchers superimposed onto the microCT scan to create a hybrid structural-functional image of PpIX fluorescence contrast. The hybrid image clearly showed evidence of the tumour, whereas the microCT image did not. Combining the two modalities does offer a clear benefit, however, as the exterior boundary seen via microCT increases the accuracy of localizing the reconstructed tumour volume.

Pogue notes that it should be possible to use fluorescence tomography to image the human brain, but that this would be slightly limited as the light cannot transmit through the entire brain. Optical tomography could, however, be used to image through parts of the head or body, or in a subsurface imaging geometry. Such subsurface imaging, currently the subject of further development, could benefit fluorescence-guided surgery by adding depth information to the procedure.

The researchers are also looking at tracking the response to antibody therapy with fluorescent molecular tracers. "This is very important, because molecular therapies are likely the wave of the future in medical oncology, and the only real hope for cancers such as glioma which have inherent microinvasive spread," Pogue told medicalphysicsweb. "If we could track the activity of molecular therapy with fluorescent tracers, then we have potential to monitor its success or failure, which could probably not be achieved with structural-based imaging."

About the author

Tami Freeman is Editor of medicalphysicsweb.