Protective hoods
What is the mantra of animal tissue culture?
Thaw quickly, freeze slowly.
Culture vessels?
Flask(25 cm2 & 75 cm2) and Petri dishes
Type of centrifuge in class?
Clinical centrifuge
DMEM?
Delbecco’s modified eagles media
Media for fibroblasts and mesenchymal cells
Cryomedia
Appropriate growth media+ 10% DMSO+ 20% serum
What are two types of hoods in ATC?
BSC I & BSC II
What are ATC vessels coated with?
Poly-l-lysine
Why is clinical centrifuge good for ATC?
Tubes swing out, allowing for a pellet to form.
mEF?
Murine embryonic fibroblast
Regular DMEM + sodium pyruvate
Powdered media has?
Serum+ sodium bicarbonate+ antibiotic/ antimyotic
BSC III protects?
Everything including hands, used for studying infectious diseases
Two types of pipettes?
Electric Pipette- controls speed of pipette
Air displacement pipette- used for cell count, cannot go above or below max and min capacity or will ruin the pipette.
Can two 4ml conical tubes and two 2 ml conical tubes be centrifuged together
Yes, as long as both 4 ml are directly across from each other and both 2 ml are directly crossed from each other. This allows them to be balanced out.
mESC?
Murine embryonic stem cell
Regular DMEM + 20% FBS & LIF+ 10% DMSO
cryopreservation
1) Mr. Frosty with isopropyl
2) add DMSO
3) Add serum
4) liquid nitrogen container
5) cryovials- where cells are stored
BSC I protects?
No protection for person using it
How can you tell if cells are contaminated?
Liquid in flask is yellow-bacterial contamination.
See bacteria swimming around in microscope.
Red cloudy
When do you centrifuge ?
Two obtain non-adherent cells, get cells that are mixed with media.
LIF?
Leukemia inhibitory factor
Enzyme keeping embryonic cells from doing what they want to do which is to create an organism
Maintain pluripotency (can become any cell) in ESC’s
Why do you thaw quickly and freeze slowly?
Prevent formation of crystals
Isopropyl helps temperature increase by 1 c
BSC II protects ?
Environment, person, and product
What is the difference between today’s flask and older flask?
Have a vented cap allowing for air exchange. Before, had to loosen cap to allow for air exchange.
What is exclusion dye?
Excludes dead cells from a count
Explant?
Chunk of tissue that is plated in hopes cells will migrate out of tissue and form a culture?
Why do we not thaw cells in incubator?
Crystals will form
Cells should be thawed in water bath 37 c