Home › Microbiology › Fabrication of Custom Agarose Wells for Cell Seeding and Tissue Ring Self-assembly Using 3D-Printed Molds
Microbiology JoVE (Open Access) Citable · DOI

Fabrication of Custom Agarose Wells for Cell Seeding and Tissue Ring Self-assembly Using 3D-Printed Molds

DOI: 10.3791/56618-v
What you'll learn
  • ✓ Design and 3D-print custom molds for variable-sized tissue ring fabrication
  • ✓ Cast PDMS negatives and agarose wells using printed mold templates
  • ✓ Seed rat aortic smooth muscle cells into agarose wells for self-assembly
Protocol

Biopharma Insights This protocol describes a platform for fabricating self-assembled tissue rings in variable sizes using a customized 3D-printed plastic mold. PDMS negatives are cured in the 3D-printed mold; then agarose is cast in the cured PDMS negatives. Cells are seeded into the resulting agarose wells where they aggregate into tissue rings.

Difficulty
intermediate
Total time
~3–4 days (including 3D printing, PDMS curing, agarose casting, and cell seeding/culture)
Model organism
Rat aortic smooth muscle cells (RaSMCs)
Biosafety
BSL-1

Steps

1
3D-print mold and cure PDMS negative

Design and 3D-print a custom plastic mold, then cast polydimethylsiloxane (PDMS) into the mold and cure according to manufacturer protocol to generate a negative replica.

▶ 00:36
2
Fabricate agarose wells from PDMS negatives

Pour molten agarose into the cured PDMS negative molds and allow to solidify, creating agarose wells of defined geometry for cell seeding.

▶ 02:41
3
Seed rat aortic smooth muscle cells into wells

Pipette rat aortic smooth muscle cells (RaSMCs) into the agarose wells and culture under conditions that promote cell aggregation into organized tissue rings.

▶ 04:01
4
Assess self-assembled tissue ring morphology

Visualize and document the resulting self-assembled tissue rings to confirm successful cell organization and ring formation.

▶ 06:56
💬 Comments coming soon