Female nmri nu nu mice

Female NMRI nu/nu mice (6–7 weeks, 20–25 g) were purchased from Charles River Laboratories, St. Germain Sur L'Arbresle, France, and received water and food ad libitum. They were kept under standardized conditions with 50 ± 10% relative humidity, 20 ± 1°C and a 12 h light-dark cycle. All animal experiments were approved by the governmental administrations of Tuebingen, Germany, and in accordance with the German Animal Protection Law.
On the day of inoculation A431 and Colo-205 cells, respectively, were trypsinized, washed twice with PBS and counted using a Neubauer chamber. 5*10⁶ A431 cells or 3*10⁶ Colo-205 cells in a volume of 200 μL PBS solution were inoculated subcutaneously into the upper right flank of the animals, respectively, while they were anaesthetized with 1.5% isoflurane vaporized in 0.8 L/min 100% oxygen.
9 days after inoculation tumors reached sizes of approximately 0.2 cubic centimeter (cc) and treatment was started after randomization of the animals. Tumor volumes were measured using calipers and calculated according to the equation: Volume[cc] = π*a*b²/6000, where a was the long and b the short axis.

Female NMRI nu/nu mice (Charles River, 12 weeks old) received s.c. injections of 3×10⁶ Colo205 cells in 100 µl PBS at the left and right dorsal sides. Treatment started 14 days after tumour cell inoculation when tumours reached an average volume of 100 mm³. 0.3 nmol HC-scTRAIL or Fc-scTRAIL-FAVSGAA were administered twice a week in 100 µl PBS for a total of six doses. The control group received six doses of 100 µl PBS. In a second round of treatment, 100 µg Smac mimetic SM83 was co-administered i.p., together with a total of 4 i.v. doses of PBS or 0.3 nmol of the proteins twice a week. Tumour volume was monitored as described^{9 (link)}. One-way ANOVA and Tukey post hoc tests were performed for statistical analysis.