Mechanical Bioreactor Final Demo Amanda Castagnino Emily Adams

Mechanical Bioreactor Final Demo Amanda Castagnino Emily Adams Emily Wood Natalie Nold Simran Singh

FINAL GOALS 1. Complete all final deliverables. FINAL DELIVERABLES 1. Assembled Device 2. Technical Paper 3. Complete EDGE Page 4. Imagine Poster 5. Full device protocol 6. MSD Documentation 7. Final Bill of Materials

Goals and Deliverables Assembled Device Software Package Performance v Requirements Cell Culture Results Next Steps Discuss handoff and leftover materials Agenda

Machining Summary ● ● Holder Device Lid Mold

Completed Device

Electrical Summary Due to the limitations for entering in selections using buttons, the following area was setup in the code for user defined variables. This will be fully explained in the protocol as well as a quick summary in the code. The students will be able to control: ● The length of the test ● The frequency of the test ● The strain for the chamber

Performance vs Requirements Mechanical and electrical systems are successful. Basic p. H may be causing poor cell viability. Longer assembly than expected due to PDMS molding and overnight cell attachment.

Experiment Process Mold PDMS at 100°C for 35 minutes. Cool. Rinse with ethanol. Rinse 3 times with PBS. Treat with ECM condition (listed on next slide). Seed cells into chamber and let attach overnight. Place cell chamber onto device and run 2 xn hour test. Image cells before and after test to see cell morphology.

Cell Culture Tests (ECM conditions) ● Condition 1: 50 μg/m. L Collagen ○ Diluted 3 mg/m. L stock with PBS ○ Incubated at room temperature 1 hr ○ Poor off matrix solution, seed cells immediately ● Condition 2: 3. 3 micrograms/m. L Fibronectin ○ Diluted 10 μg/m. L stock with PBS ○ Incubated in incubator overnight ○ Poor off matrix solution, seed cells immediately ● Condition 3: 50 micrograms/m. L Collagen ○ Diluted 3 mg/m. L stock with 20 m. M Acetic Acid ○ Incubated at room temperature 1 hr ○ Poor off matrix solution ○ Wash chamber 3 X’s with PBS solution ○ Seed cells immediately

PDMS Imaging Pre-/Post- Stretching Before

10 X - After 2 hrs 4 X - After 2 hrs 10 X - Before 10 X - Flask

Next Steps ● PDMS surface treatment ○ ○ ● Investigate p. H issues ○ ● Polish down and smooth mold for ease of imaging Brainstorm better method of transportation ○ ○ ● Look more into CO 2 diffusion with chamber design Smooth Metal Mold ○ ● Plasma treatment ~ change hydrophobic properties of the PDMS surface. Consider alternative ECM coatings/procedure. Need a smoother way to move mold in and out of incubator sterilely Want to be able to image mold in holder so mold doesn’t have to move around as much Software ○ More user friendly interface.

Bill of Materials Over $300 remaining in budget.

P 18081 Wrap-Up Gate Review - Wednesday, May 2 in the morning? Technical Paper and Performance v Requirements - to be submitted today EDGE page - to be completed by Imagine. Results pending. Final device protocol - To be updated and sent to customer next week. MSD Documents - hard copies or access to Google Drive folder? Device Storage?