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).

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