Tumor Implantation and Injection of a Tumor Vaccine in a Mouse Model

Intradermally implant 500,000 cells B16-F10 tumor cells in 50 microliters of cold PBS, and the left flank using a 30-gauge needle. The dose of implanted B16-F10 tumor cells may need to be adjusted in a range of 50,000 to 500,000 cells for successful tumor development.

After implementation, measure the tumor length and width three times a week using an electronic digital caliper. Calculate the tumor volume, and treat the mice with tumor vaccine when tumors have reached a size of 2 millimeters as described in the text manuscript.

Using an X-ray irradiator, set at 160 kilovolts and 25 milliamperes. Irradiate cells at 150 gray doses of gamma rays. Count and check the cell viability by trypan blue staining before injection. Intradermally inject the mice with one million irradiated B16-Flt3L cells in 50 microliters of cold PBS on the same flank as the original tumor implantation approximately 1 centimeter from the site of the primary tumor on days three, six, and nine after the initial cell implantation.

Mark the vaccine injection sites with a colored pen to distinguish them from the primary tumor. If 50,000 B16-F10 cells were initially implanted, it is recommended to perform vaccine treatment on days 8, 11, and 14.