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An orthotopic rat tumour recurrence model for malignant pleural mesothelioma (MPM) provides clinical similarity to patients and is useful for drug testing combined with surgical intervention. Importantly, a reliable imaging method is required allowing for noninvasive and repetitive evaluation of the tumour load. We compared the tumour load assessed by bioluminescence and magnetic resonance imaging (MRI) to the macroscopic tumour volume as a reference standard.A total of 500 000 syngeneic rat MPM cells transfected with luciferase were implanted underneath the parietal pleura of immunocompetent rats (n = 13). From the second day after implantation, bioluminescence measurements of the tumour load expressed as the maximum bioluminescent intensity (photon/second) were performed daily after intraperitoneal injection of the luciferase substrate, D-luciferin, to observe the first occurrence of tumour. Six days after the first detection of tumour, bioluminescence, MRI and macroscopic tumour volume measurement were conducted. For MRI, a 4.7-Tesla small animal imager equipped with a 1H whole-body rat coil was employed using T2-weighted fast spin-echo sequences. Tumour burden (mm3) was quantified from magnetic resonance transverse images by two independent readers by manual segmentation. Finally, the tumour burden assessed by bioluminescence and MRI was correlated (Pearson's correlation) with the macroscopic measurement of tumour (ellipsoid) volume.In all rats, a single tumour nodule was found at the inoculation site with a median macroscopic volume of 46 mm3 (18–377 mm3). For tumour burden quantification of MRIs, we observed good interobserver correlation (R2 = 0.81, P < 0.0001) as well as significant association with the macroscopic tumour volume (R2 = 0.59, P = 0.002). However, the signal intensity of bioluminescence did not correspond to the macroscopic tumour volume (R2 = 0.01, P = 0.76).MRI is a reliable and reproducible noninvasive in vivo imaging method for MPM tumour burden assessment for the present MPM model.