Purpose: Dedicated small‐animal imaging devices are being used more frequently for translational molecular imaging studies. However, few studies have investigated the magnitude of animal motion during extended dynamic imaging studies or the precision of animal repositioning in multi‐modality and/or serial imaging protocols. The objective of this work was to determine the positional accuracy and precision with which tumors in situ can be reliably and reproducibly imaged on dedicated small‐animal imaging equipment. Method and Materials: A custom rodent animal cradle with a stereotactic template served to define a coordinate system and to facilitate rigid‐body image registration. Attached to the template were fiduciary markers containing PET tracer and MRI and CT contrast media for visualization on the respective scanners. To quantify animal tumor motion during imaging protocols, “gold standard” point markers were inserted into tumors grown on the hind limb of nude rats. Three types of imaging examination were performed with the animals continuously anesthetized and immobilized: (i) single‐modality imaging (microPET and MRI) in which the animals remained in the same scanner continuously for 2 hours, (ii) multi‐modality imaging studies in which the animals were transported from a microPET to an MR scanner located in another building, and (iii) serial microPET scans in which the animals were removed from the scanner, then re‐positioned and scanned. Results: The animal tumor moved by less than 0.2–0.3 mm over two‐hour microPET or MR imaging sessions. Transporting the animal between instruments introduced additional error of ∼0.2 mm. In serial animal imaging studies, in which the animal was returned to its cage and subsequently re‐positioned, reproducibility within ∼0.8 mm could be obtained. Conclusion: To our knowledge, this is the first study systematically and rigorously evaluating the accuracy and precision with which tumors can be repeatedly imaged in small‐animal imaging devices.

Purpose: To optimize the correlation between interstitial pO2 readings and PET‐derived concentrations in tumors using co‐registered Gd‐DTPA DCE‐MRI to distinguish viable and necrotic tumor cells. Method and Materials: Nude rats with Dunning R3327‐AT xenografts were positioned in a custom‐fabricated mold and imaged by (1) T2‐weighted MRI imaging; (2) DCE‐MRI following IV injection of gadopentetate dimeglumine; (3) microPET for 1/2 hr post‐IV injection of or . The MRI and PET images were used to direct intra‐tumoral pO2 measurements using an image‐guided robot system. PET‐ and MRI‐visible fiducial markers were used to register the respective image and robot coordinate systems. pO2 was measured at 0.5‐mm increments along different tracks within the tumor using an OxyLite™ 4000 Oxygen probe advanced by the robot, providing point‐to‐point correspondence between the pO2 measurement and image‐voxel intensity. Measurement points in the necrotic region and those outside tumor region, identified by DCE‐MRI and T2‐weighted MRI, were excluded from the correlation analysis. Necrotic tissue was identified as having at t=2 min after contrast injection, where I0 and I(t) are MR image intensities before injection and at time t. Results: The registration error between images and the robot is <0.3mm. For three FMISO studies with 539 measurements and 21 tracks, the correlation between interstitial pO2 readings and intensities was not improved — vs . Four animals were studied using with 423 measurements and 20 tracks. The negative correlation improved significantly (p =0.00118) — vs — if only viable‐tumor points were considered. Conclusion: The negative correlation between voxel intensities and intra‐tumoral pO2 was improved when DCE‐MRI was used to exclude necrotic tissue pO2s. Exclusion of necrotic points did not improve the correlation for , however.

Purpose: To establish a criterion for distinguishing viable and necrotic tumor cells using the initial relative slope of the Gd‐DTPA DCE‐MRI and to apply this criterion to optimize the specificity of tumor hypoxia imaging based on PET. Method and Materials: Three nude rats with Dunning R3327‐AT prostate adenocarcinoma xenografts were imaged by dynamic MRI following tail vein injection of gadopentetate dimeglumine, with imaging parameters of 1‐mm slice thickness, 0.5‐mm spacing, and 0.13mm × 0.13mm voxel size. The time‐intensity curve of each voxel was obtained at 1‐min interval to 25‐min post‐injection. Histologic (standard H&E stain and high‐power microscopy) examination was performed of 8‐μm thick slices parallel to the DCE‐MRI imaging axis and the necrotic regions were identified. The initial relative slope, where I 0 and I(t) are MR image intensity before injection and at time t, was calculated for different times and compared with the pathologically defined necrotic region to determine the threshold that best distinguishes viable and necrotic cells. This criterion was then used to study the specificity of FMISO PET with the same xenografts scanned with DCE‐MRI and PET. Results: The optimal criterion for identifying viable and necrotic tumor regions was that a DCE‐MRI voxel was necrotic if at 2 minutes after contrast injection. When this criterion was applied to the PET images of three xenografts, necrotic region was found to have a wide range of image intensities, which is inconsistent with the hypothesis that high image intensity exclusively identifies hypoxic viable tumor. Among the 319 hypoxic voxels determined from the three animals' PET, only 75% corresponded to viable cells. Conclusion: A criterion was established to identify necrotic/viable tumor cells using DCE‐MRI. Using PET alone for hypoxia imaging is problematic because the specificity might be compromised by necrotic regions with high PET image intensity.