Cell lines

The tumor cell line WM115 and SK-MEL-2 were obtained from ATCC between 2016 and 2018, whereas M14 cells were provided by Dr. Ferrone (Massachusetts General Hospital). The tumor cell lines MDA-MB-468 and MDA-MB-231 were obtained from German Collection of Microorganism and Cell Cultures GmbH (DSMZ, ACC 738 and ACC 732). The 293T cells used for the production of retroviral vectors were obtained from ATCC in early 2000. All cells were maintained in culture with the appropriate media, either RPMI-1640 (Gibco), DMEM (Gibco), or MEM (Gibco) supplemented with 10% FBS (Sigma), 1% L-glutamine (Gibco), and 1% penicillin/streptomycin (Gibco) in a humidified atmosphere containing 5% CO₂ at 37°C. WM115 cells were modified to express the fusion protein firefly luciferase and enhanced GFP (eGFP-FFluc)(11). Cells were kept in culture for less than six consecutive months, after which, aliquots from the original expanded vial (cultured for 4 to 6 passages after being received) were used. All tumor cell lines were routinely tested to exclude contamination with Mycoplasma and assessed for the expression of transgenes and tumor markers by flow cytometry to confirm identity. Glioblastoma-derived neurospheres (GBM-NS) were generated as previously described (12). The variable regions of the immunoglobulin genes expressed by the 763.74 hybridoma were amplified with a set of proprietary primers from cDNA generated from the hybridoma cells using a RT-PCR protocol and sequenced using a standard dye-terminator capillary sequencing method (SynBuild, Tempe, Az).