Robust tools for the quantitation of perfusion are not fully developed using contrast enhanced ultrasound (CEUS). The ovine corpus luteum (CL) is a transient gland in the ovary that is formed to produce the hormone progesterone essential for maintenance of pregnancy. Importantly, it has a dense microvascular network with predictable and well-regulated angiogenic mechanisms. In a number of different experiments it was shown that this property may be used to investigate and refine imaging methodology. Using a Philips iU22 ultrasound scanner (Philips Medical Systems Corp, Seattle, WA) in contrast imaging mode it was shown that a highly controlled experiment may produce high levels of reproducibility in the transit of contrast with standard uncertainty below 10%. Also, compartmental kinetics models were tested. The use of prostaglandin F2alpha promotes an intense anti-angiogenesis, allowing monitoring with CEUS prior to and following the demise of the CL microvasculature within 24 hours. Finally, the robust angiogenic property of the CL during the oestrous cycle allows further refinement of CEUS in vivo. In conclusion, the CL offers an attractive changing vascular bed for assessing existing and developing new clinically relevant perfusion imaging methodology.