*4.5. PET Imaging Procedure*

For PET imaging, mice were pre-anesthetized in an induction chamber using isoflurane and then positioned in a prone position on a double imaging chamber (m2m Imaging Corp, Cleveland, OH, USA). Two mice were imaged simultaneously during one PET acquisition. Animals were warmed throughout the experiment and body temperature and respiratory rate were constantly monitored (SA Instruments Inc, Stony Brook, NY, USA). The level of isoflurane concentration was adjusted (range 1.5–3% in air) during the imaging procedure to achieve a constant level of anesthesia. The imaging chamber was positioned in the gantry of a microPET scanner (Focus 220, Siemens Medical Solutions, Knoxville, TN, USA) and a 10 min transmission scan using a rotating <sup>57</sup>Co point source was recorded. Subsequently, 60-min dynamic emission scans (energy window 250–750 keV; timing window = 6 ns) were started with the injection of either [11C]tariquidar (25 <sup>±</sup> 7 MBq) or [11C]erlotinib (22 <sup>±</sup> 5 MBq) injected i.v. at a volume of 0.1 mL over a period of 120 s.

After completion of the PET scan, a blood sample (20–30 µL) was collected from the retro-orbital venous plexus, the mice were sacrificed by cervical dislocation and transcardially perfused using 10 mL phosphate-buffered saline. Blood was centrifuged (13,000× *g*, 4 ◦C, 4 min) to obtain plasma and whole brains were removed and processed for immunohistochemistry as described below. Aliquots of blood and plasma were transferred into pre-weighted test tubes and measured in a gamma-counter (HIDEX AMG Automatic Gamma Counter, Turku, Finland). Filled tubes were weighed to obtain tissue weight. The gamma-counter was calibrated using a series of tubes with decreasing activity of a <sup>11</sup>C-solution. The measured radioactivity data were decay-corrected to the time of radiotracer injection and expressed as standardized uptake value (SUV), which is calculated as follows: (radioactivity per g (kBq/g)/injected radioactivity (kBq)) × body weight (g).
