Concordance between alizarin red stained skeleton and micro‐CT skeleton evaluation methods: A case study in New Zealand White rabbits

Abstract

ObjectivesThe objective of this study was to examine the fetal skeletons using both alizarin red stain and micro‐computed tomography (CT) images; investigate differences, and to determine if the conclusions of the study were the same regardless of the examination method.MethodsA candidate drug was given orally by gavage to pregnant New Zealand White rabbits on gestation day (GD) 7 to GD 19 (mating = GD 0) at doses of 0 (control), 0.02, 0.5, 5, and 15 mg/kg/day. Maternal toxicity was evident at ≥0.02 mg/kg/day. The 199 fetal skeletons (totaling 50,546 skeletal elements) obtained at cesarean delivery on GD29 were first stained with Alizarin Red S, then imaged by a Siemens Inveon micro‐CT scanner. All fetal skeletons were examined by both methods, without knowledge of dose group, and the results were compared.ResultsIn total, 33 types of skeletal abnormalities were identified. There was 99.8% concordance of results comparing stain to micro‐CT. Ossification of the middle phalanx of the forepaw digit 5 showed the greatest difference between the two methods.ConclusionOverall, micro‐CT imaging is a realistic, and robust alternative to skeletal staining to examine fetal rabbit skeletons in developmental toxicity studies.