Figure 1.

Illustration of the set-up for the in vivo experiment. A nude mouse with a xenograft prostate tumor growing on the flank was intraperitoneally injected with both FAZA and pimonidazole. Approximately four hours later, the mouse was sacrificed, and the tumor was excised and fresh frozen. The tumor was immediately cryo-sectioned in three layers, each layer consisting of 10 consecutive 10 μm sections. Then, these sections were stained for either hematoxylin and eosin (HE) or pimonidazole binding, or used for FAZA autoradiography. The autoradiogram was used as a template for laser micro dissection, where the hypoxic (H) and the non-hypoxic (NH) areas were identified and cut. HE staining was used to evaluate and avoid necrotic areas. This dissected tissue was immediately lysed and real-time PCR (qPCR) performed. The images of the laser micro dissection technique are from Arcturus Bioscience (Mountain View, CA, USA).