Generating an Organoid Culture from a Brain Tumor Cell Suspension

Begin with a microcentrifuge tube containing a laminin-rich extracellular matrix or lrECM. Laminin is a structural component present abundantly in the brain.

Keep the tube on ice to prevent IrECM polymerization.

Add brain tumor cells and mix thoroughly to ensure a uniform distribution of cells.

Next, transfer the mixture onto parafilm molds, forming droplets.

Incubate to facilitate lrECM polymerization to form a scaffold, mimicking the cells' natural environment.

Now, flush the solidified organoids into a Petri dish containing a culture medium, then incubate.

The culture medium provides essential nutrients and growth factors, supporting cell division.

Within the organoid, cells self-organize by migrating and invading.

After incubation, tilt the plate to settle the organoids.

Carefully remove the excess medium and add fresh medium.

Incubate with gentle shaking.

This distributes the nutrients evenly and enhances organoid growth.

To make organoids from a single-cell suspension, add an appropriate amount of laminin-rich extracellular matrix or lrECM into a small centrifuge tube in an ice bucket or cold block.

Prepare a cell mixture containing 20,000 cells per organoid. Mix the lrECM ce