April 1, 2014
Objectives
Au:Myoglobin Nanofibers: Sample Prep for Cell Culture
- Solutions:
- TRIS dopamine: 0.0475g dopamine hydrochloride and 0.03g TRIS in 25mL to make 10mM solution at pH 8.46
- Au: 0.011g in 10mL to make 2.793mM solution
- Myoglobin: 0.011g in 10mL to make 0.0621mM solution
Plain Scaffolds
- 4 PLA scaffolds printed for each test tube
- Three test tubes filled with scaffolds and water
- Autoclaved
Brute Force Nanofiber-coated Scaffolds
- 4 PLA scaffolds printed for each test tube
- Gold:Myoglobin (100:1) nanofibers synthesized by above method
- Nanofibers dried onto PLA scaffolds
- Scaffolds submerged in water
- Autoclaved
Original Method for Nanofiber-coated Scaffolds
- 4 PLA scaffolds printed for each test tube
- Gold:Myoglobin (100:1) nanofibers synthesized by above procedure with scaffolds in solution
- Autoclaved
Polydopamine-coated Scaffolds
- 4 PLA scaffolds printed for each test tube
- Scaffolds submerged in 10mM pH 8.46 TRIS dopamine hydrochloride solution for 1 hour
- Add 1mL of 0.0621 Myoglobin to each test tube, shake tube to combine, allow surface functionalization to occur for 5 hours
- Rinse with deionized water and allow scaffold to dry overnight
- Scaffolds submerged in water
- Autoclaved
Polydopamine Nanofiber-coated Scaffolds
- 4 PLA scaffolds printed for each test tube
- Scaffolds submerged in 10mM pH 8.5 TRIS dopamine hydrochloride solution for 1 hour
- Add 1mL of 0.0621 Myoglobin to each test tube, shake tube to combine, allow surface functionalization to occur for 5 hours
- Rinse with deionized water and allow scaffold to dry overnight
- Gold:Myoglobin (100:1) nanofibers synthesized by above procedure with polydopamine-coated scaffolds in solution
- Autoclaved
Breakdown
Today
- Time sensitive: Make 40mL TRIS-polydopamine solution
- Time sensitive: Make 6 tests tubes of TRIS-polydopamine solution, put scaffolds in all 6 test tubes at 12 p.m. (3 for pD only, 3 for pD-original)
- Make Au and Myoglobin solutions
- Make 9 test tubes of 100:1 Au:Myo Nanofibers (3 for original, 3 for brute force fibers, 3 for pD-original tomorrow; see volumes in chart above)
- Time sensitive: At 1 p.m. add 1mL of 0.0621M Myoglobin to all 6 tubes (3 for pD only, 3 for pD-original)
- Put scaffolds into 3 of the 9 Au:Myo test tubes when finished (3 for original)
- Make 3 test tubes of water -> put scaffolds into 3 test tubes when finished
- Put 9 test tubes (3 brute force, 3 original, 3 water) into oven, set for 4 hours at 80C
- Time Sensitive: At 6 p.m. rinse pD scaffolds that have soaked for 5 hours, submerge 6 in water overnight (if possible, submerge 3 of 6 tubes in Au:Myo solution and put all 6 in oven for 4 hours), dump Au:Myo fibers onto scaffolds for brute force
Tomorrow
- If oven was not finished yesterday: submerge 3 pD scaffolds into original solution, leave other 3 in water and put in oven for 4 hours at 80C
- Submerged dried brute force scaffolds in water
- Autoclave all: 3 plain, 3 brute force, 3 original, 3 pD-Myo, 3 pD-Myo-Au
Type I Collagen Test (Detection and Quantification)
Collagen Staining Procedure & Microscope Imaging
Trypsin Cell Detachment Procedure & Cell Counting
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Cell/Scaffold Tests
