Differentiation of 3T3-L1 Cells: Difference between revisions

Line 6:

*'''NCS''' (currently we are using Biowhittaker Cambrex cat# 14-416F, lot# 01106776; aliquot in 50 mL tubes and store at -20)

*'''PSG''' (100X Invitrogen Cat# 100378-016)

*'''Insulin''' (Sigma I-5523). Dissolve in 0.01N Hydrochloric acid as a 1 mg/ml stock. Aliquot into 1.5 mL tubes and store at -20

*'''Insulin''' (Sigma I-5523). Dissolve in 0.01N Hydrochloric acid (28.7μl/50ml) as a 1 mg/ml stock. Aliquot into 1.5 mL tubes and store at -20

*'''Dexamethasone''' (Sigma D-1756). Dissolve in ethanol as a 2.5 mM stock. Aliquot into 1.5 tubes and store at -20.

*'''Dexamethasone''' (Sigma D-1756). Dissolve in ethanol as a 2.5 mM stock (0.049g/50ml). Aliquot into 1.5 tubes and store at -20.

*'''MIX''' (Sigma I-5879). Dissolve 2.78g/50mL KOH (0.98g/50mL). Aliquot into 1.5 mL tubes and store at -20

*'''Fibroblast Media''' (DMEM + 10% NCS + 1X PSG; Sterile filter into new bottle)

*'''Adipocyte Media''' (DMEM + 10 % FBS + 1X PSG; Sterile filter into new bottle)

*'''Adipocyte Media''' (DMEM + 10 % L1-FBS + 1X PSG; Sterile filter into new bottle)

*'''DMI Media''' (Adipocyte Media + Insulin (1000X dilution), Dexamethasone (10 000X dilution) and MIX (500X dilution); Sterile filter into new bottle after additions

*'''Insulin Media''' (Adipocyte Media + Insulin (1000X dilution); Sterile filter into new bottle after adding insulin)

@@ Line 6: / Line 6: @@
 *'''NCS''' (currently we are using Biowhittaker Cambrex cat# 14-416F, lot# 01106776; aliquot in 50 mL tubes and store at -20)
 *'''PSG''' (100X Invitrogen Cat# 100378-016)
-*'''Insulin''' (Sigma I-5523).  Dissolve in 0.01N Hydrochloric acid as a 1 mg/ml stock.  Aliquot into 1.5 mL tubes and store at -20
+*'''Insulin''' (Sigma I-5523).  Dissolve in 0.01N Hydrochloric acid (28.7μl/50ml) as a 1 mg/ml stock.  Aliquot into 1.5 mL tubes and store at -20
-*'''Dexamethasone''' (Sigma D-1756).  Dissolve in ethanol as a 2.5 mM stock.  Aliquot into 1.5 tubes and store at -20.
+*'''Dexamethasone''' (Sigma D-1756).  Dissolve in ethanol as a 2.5 mM stock (0.049g/50ml).  Aliquot into 1.5 tubes and store at -20.
 *'''MIX''' (Sigma I-5879).  Dissolve 2.78g/50mL KOH (0.98g/50mL).   Aliquot into 1.5 mL tubes and store at -20
 *'''Fibroblast Media''' (DMEM + 10% NCS + 1X PSG; Sterile filter into new bottle)
-*'''Adipocyte Media''' (DMEM + 10 % FBS + 1X PSG; Sterile filter into new bottle)
+*'''Adipocyte Media''' (DMEM + 10 % L1-FBS + 1X PSG; Sterile filter into new bottle)
 *'''DMI Media''' (Adipocyte Media + Insulin (1000X dilution), Dexamethasone (10 000X dilution) and MIX (500X dilution); Sterile filter into new bottle after additions
 *'''Insulin Media''' (Adipocyte Media + Insulin (1000X dilution); Sterile filter into new bottle after adding insulin)