2.2. U87 Glioblastoma Cell Line Culture and Transplantation

The green fluorescent protein (GFP)-transfected human GB cell line U87 (U87GFP+) was obtained from ATCC/LGC (Wesel, Germany). Cells were maintained at a temperature of 37 °C in an atmosphere of 5% CO₂/95% air with Dulbecco’s modified Eagle’s medium (DMEM; Biochrom, Berlin, Germany) supplemented by 10% fetal bovine serum (FBS, Biochrom) and 1% penicillin/streptomycin (Biochrom). Cells were detached from the culture flasks using 0.05% trypsin/ethylenediaminetetraacetic acid (EDTA) before tumor transplantation.

On the day after OBSC preparation, 10,000 U87 cells in 0.1 µL medium were transplanted bilaterally onto the CA3 region of the hippocampus. The hippocampus is together with the subventricular zone a germinative niche in the mammalian brain, both vulnerable sites for the growth of transformed cells, because they are abundant in growth factors and permissive to proliferation [18]. The treatment started seven days after the OBSC preparation (for timeline see Figure 1A).

Experimental design. (A) Timeline. U87GFP+ cells (10,000 cells/0.1 µL medium) were implanted bilaterally into the CA3 region of the hippocampus 24 h after organotypic brain slice culture preparation. One intersection represents 24 h resp. 1 day. (B) Quantification of the tumor area.