Fall 2015Amelia Payne | Big Picture South Burlington
Introduction
• Feel free to use electronic devices during my presentation
• Ask questions during presentation
• You will get candy
Introduction
• Internship
• Classes
• Online Learning
• Time tracking
• Clubs
• STP
Internship At Weiss Lab
Freezing Decellurized Lungs
• Cultivate cells in sterile conditions
• Freeze tissue in different mediums
• Defrost tissues and seed with cells
• Test for cell growth with WST-assay
Growing Cell Lines Human Lung Fibroblasts (HLF)
• Defrosted cells
• Grew under sterile conditions
• Counted
• Replated and frozen
Counting
• Inject 10 µl of cell suspension into “Neubauer chamber” (device for cell counting)
• Count number of cells in one quadrant
• Multiply by 10^4 to get total cells per ml cell suspension
Freezing Lungs• At rate of 1°C per minute
to -80°C
• Transferred to liquid nitrogen (-195°C = -320°F) for long term storage
• Will defrost at 1, 3, 6, and 12 months and do WST-assay
• Control: fresh tissue
• We froze tissue in
• 5% glycerol
• 10 % DMSO in PBS
Seeding Defrosted Tissue
We will defrost tissue rapidly and seed with cells once we reach the first time points.
WST-assay
• Indicates metabolic activity of cells
• WST-1 (water soluble tertrazolium salt-1, red) is cleaved into formazan (yellow) by an enzyme produced by the mitochondria. The resultant color changes indicates metabolic activity of the cells
• Absorbance measurement (440 nm) to determine how much formazan was produces
Data from WST Assay Control
The Team
Thanks to all my awesome mentors who are here today
*Sorry if you’re not in the picture because you weren't at lunch on Thursday.
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