Tissue vasculature is essential to maintaining its viability and functionality, however, there are currently no simple ways to create thick vascularized tissue. This protocol enables researchers to create thick vascularized tissue cast in a hydrogel with any vascular network design. The tissue unit created with this kit can be easily connected to other tissue units and pumps.
You Will Need
- Allevi vascularization chip
- 3 mL of pluronic
- 1.5 mL 20% w/v GelMA
- 0.5 g LAP Photoinitiator
- Allevi 5 mL syringe
- 2 x 25G tapered plastic needles
- 5 x 30G plastic needles
- Syringe coupler
- Cell media
- Follow this protocol to prepare your GelMA with cells (or this protocol for lyophilized gelMA prep);
- Attach the 25G plastic tapered needle to the GelMA syringe and place it in the left extruder;
- Attach the 30G plastic tip to the pluronic syringe and load the right extruder;
- NOTE: You may use lower gauges for thicker channels.
- Set the left extruder temperature to 30˚C;
- Wait for about 5 minutes until the GelMA solution reaches 30˚C;
- Use the grid on the gasket slide to calibrate the right extruder to the x, y and z coordinates that align with the gasket inlet and outlet;
- Calibrate the right extruder at these coordinates;
- Calibrate the left extruder at the bottom right corner of the gasket, with the tip right at the top surface;
- Use our straight line or convoluted channel file, which codes for crosslinking, or create your own file that:
- Extrudes GelMA at 10 PSI for 4 s, crosslinks GelMA for 60s at 10 mW/cm², prints the desired pluronic channel at 100 PSI and 5 mm/s, extrudes GelMA at 10 PSI for 4 s and crosslinks it for 60s at 10 mW/cm².
- Note: the total gasket volume is 3 mL and the dimensions are 1.6 x 1.6 x 1.1 cm.
- After crosslinking, place your gasket in 4˚C for 5 minutes;
- Connect a pump or syringe to the luer lock gasket connection and perfuse the channel with cold (4˚C) media until all of the pluronic is washed away;
- Culture your vascularized tissue as desired.
|Speed (mm/s)||Layer height (mm)||Nozzle Diam (mm)||Gauge|
|Pressure (PSI)||Crosslink (sec)||Print Temp (°C)|
Kolesky, D. B., Truby, R. L., Gladman, A. S., Busbee, T. A., Homan, K. A., & Lewis, J. A. (2014). 3D Bioprinting of Vascularized, Heterogeneous Cell-Laden Tissue Constructs. Advanced Materials, 26(19), 3124–3130. https://doi.org/10.1002/adma.201305506