Co-Culturing Glioblastoma Cells on Patterned Neurons: A Technique to Monitor the Migration of Glioblastoma Cells on Rat Hippocampal Neurons In Vitro
Published: April 30, 2023
Abstract
Source: Guyon, J. et al. Co-culture of Glioblastoma Stem-like Cells on Patterned Neurons to Study Migration and Cellular Interactions. J. Vis. Exp. (2021)
This video describes a co-culture assay of glioblastoma cells on micropatterned neurons, which mimic the myelinated neuronal tracts invasion by the glioblastoma stem-like cells as they migrate on neurons in the peritumoral spaces. This assay can help analyze the effects of pharmacological drugs that inhibit glioblastoma cells' migration on neurons in a glioma.
Protocol
All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board. 1. Preparation of the Patterned Slides Substrate Preparation for Micropatterning Treat 18 mm circular glass coverslips by air/plasma activation for 5 min. Place the coverslips in a closed chamber with 100 μL of (3-aminopropyl) triethoxysilane in a desiccator for 1 h. Incubate wi…
Açıklamalar
The authors have nothing to disclose.
Materials
| (3-aminopropyl) triethoxysilane | Sigma | 440140-100ML | The amino group is useful for the bioconjugation of mPEG-SVA |
| 96-well round-bottom plate | Sarstedt | 2582624 | Used to prepare spheroids |
| Accutase | Gibco | A11105-01 | Stored at -20 °C (long-term) or 4°C (short-term), sphere dissociation enzyme |
| B27 | Gibco | 12587 | Stored at -20 °C, defrost before use |
| Basic Fibroblast Growth Factor | Peprotech | 100-18B | Stored at -20 °C, defrost before use |
| Countess Cell Counting Chamber Slides | Invitrogen | C10283 | Used to cell counting |
| Coverslips | Marienfeld | 111580 | Cell culture substrate |
| Dessicator cartridges | Sigma | Z363456-6EA | Used to reduce moisture during (3-aminopropyl) triethoxysilane treatment |
| DPBS 10x | Pan Biotech | P04-53-500 | Stored at 4 °C |
| Flask 75 cm² | Falcon | 10497302 | |
| HBSS | Sigma | H8264-500ML | |
| Heparin sodium | Sigma | H3149-100KU | Stored at 4 °C |
| Laminin | 114956-81-9 | Promotes neuronal adhesion | |
| Flask 75 cm² | Falcon | 10497302 | |
| Heparin sodium | Sigma | H3149-100KU | Stored at 4 °C |
| Leonardo software | Loading of envisioned micropatterns | ||
| MetaMorph Software | Molecular Devices LLC | NA | Microscopy automation software |
| Methylcellulose | Sigma | M0512 | Diluted in NBM for a 2% final concentration |
| Neurobasal medium | Gibco | 21103-049 | Stored at 4 °C |
| Nikon TiE (S Fluor, 20x/0.75 NA) | Inverted microscope equipped with a motorized stage | ||
| Penicillin – Streptomycin | Gibco | 15140-122 | Stored at 4 °C |
| PLPP | Alveole | Photoinitiator used to degrade the PEG brush | |
| Poly(ethylene glycol)-Succinimidyl Valerate (mPEG-SVA) | Laysan Bio | VA-PEG-VA-5000-5g | Used as an anti-fouling coating |
| Trypsin-EDTA | Sigma | T4049-100ML | Used to detach adherent cells |
Etiketler
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Co-Culturing Glioblastoma Cells on Patterned Neurons: A Technique to Monitor the Migration of Glioblastoma Cells on Rat Hippocampal Neurons In Vitro. J. Vis. Exp. (Pending Publication), e20680, doi: (2023).