Differentiation of 3T3-L1 Cells: Difference between revisions

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==Materials==

*DMEM (4.5 g/L glucose + L-glutamine – sodium pyruvate; Invitrogen catalog # 11965-092)

*'''DMEM''' (4.5 g/L glucose + L-glutamine – sodium pyruvate; Invitrogen catalog # 11965-092)

*FBS (we test several batches from several sources for differentiation and insulin stimulated glucose uptake then order large quantities of that particular lot. Currently we are using Sigma cat# F2442, lot# 072K8403. Aliquot and store in 50 mL tubes at -20)

*'''FBS''' (we test several batches from several sources for differentiation and insulin stimulated glucose uptake then order large quantities of that particular lot. Currently we are using Sigma cat# F2442, lot# 072K8403. Aliquot and store in 50 mL tubes at -20)

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

*'''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)

*'''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 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. 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

*'''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)

*'''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 % FBS + 1X PSG; Sterile filter into new bottle)

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

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

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

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

==Fibroblast Culture==

@@ Line 1: / Line 1: @@
 ==Materials==
-*DMEM (4.5 g/L glucose + L-glutamine – sodium pyruvate; Invitrogen catalog #  11965-092)
+*'''DMEM''' (4.5 g/L glucose + L-glutamine – sodium pyruvate; Invitrogen catalog #  11965-092)
-*FBS (we test several batches from several sources for differentiation and insulin stimulated glucose uptake then order large quantities of that particular lot.  Currently we are using Sigma cat# F2442, lot# 072K8403.  Aliquot and store in 50 mL tubes at -20)
+*'''FBS''' (we test several batches from several sources for differentiation and insulin stimulated glucose uptake then order large quantities of that particular lot.  Currently we are using Sigma cat# F2442, lot# 072K8403.  Aliquot and store in 50 mL tubes at -20)
-*NCS (currently we are using Biowhittaker Cambrex cat# 14-416F, lot# 01106776; aliquot in 50 mL tubes and store at -20)
+*'''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)
+*'''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 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.  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
+*'''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)
+*'''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 % FBS + 1X PSG; Sterile filter into new bottle)
-*Differentiation Media I (Adipocyte Media + Insulin (1000X dilution), Dexamethasone (10 000X dilution) and MIX (500X dilution); Sterile filter into new bottle after additions
+*'''Differentiation Media I''' (Adipocyte Media + Insulin (1000X dilution), Dexamethasone (10 000X dilution) and MIX (500X dilution); Sterile filter into new bottle after additions
-*Differentiation Media II (Adipocyte Media + Insulin (1000X dilution); Sterile filter into new bottle after adding insulin)
+*'''Differentiation Media II''' (Adipocyte Media + Insulin (1000X dilution); Sterile filter into new bottle after adding insulin)
 ==Fibroblast Culture==