20.109(S07): TA's notes for module 2: Difference between revisions
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===Notes:M2D2=== | ===Notes:M2D2=== | ||
#Need lab quiz. | #Need lab quiz. | ||
#Need 6 boxes of cuvettes | |||
#Need 12 96 well plates | |||
#Will need 6 flasks of MES cells/lab section as well as pre-transformation media for plating on slides. | #Will need 6 flasks of MES cells/lab section as well as pre-transformation media for plating on slides. | ||
#Will need at least 12 Lab-Tek slides per section. | #Will need at least 12 Lab-Tek slides per section. | ||
===Notes:M2D3=== | ===Notes:M2D3=== | ||
===Notes:M2D4=== | ===Notes:M2D4=== |
Revision as of 12:44, 19 January 2007
Ca Sensor Module
General notes
To get MES cell culture ready
Sunday | Monday | Tuesday | Wednesday | Thursday | Friday | Saturday |
one week before first day have 2 confluent flasks | ||||||
Day 1 of module | Day 2 of module | |||||
Day 3 of module | Day 4 of module |
Daily Notes
Notes:M2D1
- No quiz needed
- Will need 14 flasks of MES cells/lab as well as appropriate media for splitting cells. If more than 12 students are in lab section, add additional flask for each additional student.
- Cells for microscope refresher can be from demo of trypsinization for first group on scopes and from students own trypsinized cells for second group on scopes.
Notes:M2D2
- Need lab quiz.
- Need 6 boxes of cuvettes
- Need 12 96 well plates
- Will need 6 flasks of MES cells/lab section as well as pre-transformation media for plating on slides.
- Will need at least 12 Lab-Tek slides per section.
Notes:M2D3
Notes:M2D4
Notes:M2D5
Notes:M2D6
Recipes/Reagents
- Growth Media: 500 ml DMEM (high glucose), 50 ml serum (FBS from Atlanta Biol), 5 ml P/S/G, 1 ml BME, 5 ml NEAA. Filter then add 50 ul LIF. Store 4°
- Pre-Tx’n Media: 500 ml DMEM (high glucose), 50 ml serum (FBS from Atlanta Biol), 5 ml 100XG, 1 ml BME, 5 ml NEAA. Filter then add 50 ul LIF. Store 4°
- 0.1% gelatin for TC dishes >10’ before adding cells: 1g/L, Heat slightly to dissolve. Autoclave. Store 4°
- Cyproheptadine hydrochloride (C6022); Working conc should be 0.1uM. Make stock of 0.1M by dissolving 100 mg bottle in 3 ml DMSO (although according to Sigma this is water soluble, I had trouble dissolving this in water). When using this stock dilute 1:1000 then final concentration of 1:1000 on cells. Store -20°
- A23187 Calcium Ionophore (C9275). Working conc should be 2 uM. Make stock of 2 mM by dissolving 5 mg bottle in 4.6 ml DMSO. Use at 1:1000 on cells. Store -20°
- BAPTA-AM (A1076). Working conc should be 30 uM. Make stock of 30 mM by dissolving 25 mg bottle in 1 ml DMSO. Use at 1:1000 on cells. . Store -20°
- CaCl2*2H20 (Mallinckrodt 4160). Working conc should be 30mM. Make stock of 3M by dissolving 4.41g in 10 ml H20. Filter sterilize. Use at 1:100 on cells. Store -20°
- EGTA (JTBaker L657-05) Working conc should be 0.5mM. Make stock of 0.5M by dissolving 1.9 g in 9 ml H20 + 1 ml 10N NaOH. Filter sterilize. Use at 1:1000 on cells. Store -20°
- CaCl2 (no H20) for std curve: 55 mg/50 ml H20 gives 10mM solution.