Primary human glioma cells (HCM-BROD-0002-C71, ATCC, Manassas, VA, USA) were cultured using conditioned medium (256–100 Propagenix Inc, Gaithersburg, MD, USA). Human adult astrocytes (Cell Applications, Inc., San Diego, CA, USA) were cultured with human astrocyte growth medium (Cell Applications, Inc., San Diego, CA, USA). Human fetal astrocytes (ScienCell Research Laboratories, Carlsbad, CA, USA) were cultured using astrocyte medium (ScienCell Research Laboratories, Carlsbad, CA, USA).
Type I collagen, extracted from bovine Achilles tendon (Stroots Locker, Goddard, KS, USA), was dissolved in acetic acid to prepare collagen solutions of 3 mg/mL, 5 mg/mL, and 10 mg/mL. To generate the 3D cell cultures, glioma cells, human adult astrocytes, and Human fetal astrocytes were seeded inside the collagen hydrogels. Collagen solution (400 µL) in the wells of a 48-well plate was neutralized with NaOH solution (1 M) and phosphate-buffered saline (PBS) solution (10× [12 (link)]. Then, 40,000 cells of each cell type were mixed with the neutralized collagen solution. Additional pre-warmed medium was then gently added into the wells with the collagen gel and cell mixture. The cell culture plate was then transferred to an incubator at 37 °C with 5% CO2.
Type I collagen, extracted from bovine Achilles tendon (Stroots Locker, Goddard, KS, USA), was dissolved in acetic acid to prepare collagen solutions of 3 mg/mL, 5 mg/mL, and 10 mg/mL. To generate the 3D cell cultures, glioma cells, human adult astrocytes, and Human fetal astrocytes were seeded inside the collagen hydrogels. Collagen solution (400 µL) in the wells of a 48-well plate was neutralized with NaOH solution (1 M) and phosphate-buffered saline (PBS) solution (10× [12 (link)]. Then, 40,000 cells of each cell type were mixed with the neutralized collagen solution. Additional pre-warmed medium was then gently added into the wells with the collagen gel and cell mixture. The cell culture plate was then transferred to an incubator at 37 °C with 5% CO2.