03/20/15
- PAGE - glucagon & insulin (for Gbox demo next week)
PAGE - glucagon & insulin
> From David Barclay's pancreas cell project
Sample prep
- Loading vol. 20 uL = 5.0 μL 4x Novagen sample buffer (blue) + 1.0 μL 1M DTT + 14 uL protein
- Dilutions - make enough for at least two lanes plus pipetting error (x3)
Protein |
Final mass |
Stock μL (x3) |
dH2O μL (x3)
|
Glucagon |
10.0 μg |
10.0 (30.0) |
4.0 (12.0)
|
Glucagon |
1.0 μg |
1.0 (3.0) |
13.0 (39.0)
|
Glucagon |
0.1 μg |
0.1 (0.3) |
13.9 (41.7)
|
Insulin* |
10.0 μg |
10.0 (30.0) |
4.0 (12.0)
|
Insulin* |
1.0 μg |
1.0 (3.0) |
13.0 (39.0)
|
Insulin* |
0.1 μg |
0.1 (0.3) |
13.9 (41.7)
|
- For insulin, start with a 1/25 dilution of the 25 mg/mL stock
- 4x sample buffer batch (x15) = 75.0 4x sample buffer + 15.0 1M DTT
> Gel lanes (for both gels)
- Page Ruler plus (15 uL)
- Glucagon 10.0 μg
- Glucagon 1.0 μg
- Glucagon 0.1 μg
- Insulin 10.0 μg
- Insulin 1.0 μg
- Insulin 0.1 μg
- Page Ruler plus (15 uL)
- Glucagon 10.0 μg
- Glucagon 1.0 μg
- Insulin 10.0 μg
- Insulin 1.0 μg
Reagent |
Volume
|
reagent 1 |
# μL
|
reagent 2 |
#
|
reagent 3 |
#
|
reagent 4 |
#
|
dH2O |
#
|
|
# μL
|
--> Reaction conditions
|