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Insulin Sensitivity Test
Materials
- 10 µl stock insulin diluted in 10 ml sterile water
-Insulin we use: 10mL vial of Lilly Humulin R Regular, insulin human injection, USP. U-100 (100 units per mL)
- OneTouch® Ultra Blood Glucose Monitoring System and test strips
- 1.0 cc insulin syringes
- 25G1/2 needle
- Stopwatch
- Scale
Method
- Mice are fasted for 6 hours by removal to a clean cage without food at the end of their dark (feeding) cycle. (Fast time: 7am-1pm.)
- Mice are weighed.
- After 6 hours of fasting, a fasting glucose level is obtained from venous blood from a small tail nick. It is measured with the OneTouch® Ultra Blood Glucose Monitoring System and test strips.
- 1.0mU/g of insulin is injected intraperitoneally.
- Blood glucose values are obtained at 15, 30, 60 and 120 minutes.
- When mice’s blood glucose levels got below 30 mg/dL mice was injected with 1mg/g of glucose.
*Tips: If the IPGTT test shows that mice are very glucose tolerant fasting time can be reduced to 4 hours or dosage should be reduced to 0.5 mU/g. If mouse’s blood glucose level drops below 30mg/dl, glucose should be injected into mouse to recover it. Always have glucose solution nearby during this test. Extra time points may be added to monitor mice’s blood glucose level to make sure it doesn’t drop below 30mg/dl. You can keep track of when mice get injected by writing down the time when it was injected. 15 mice should be the max for this test.
Calculating Dosage:
Since insulin concentration is so little to inject we dilute it with sterile H2O or PBS.
Determining dose amount:
Stock Humulin insulin: 100U/ml (Always check for expired date)
Insulin dosage: 1.0 mU/g
Dilution: 10ul stock insulin in 10 ml PBS or H2O
1. Calculate dilution concentration of insulin:
100U/ml x 1ml/1000µl x 10µl/10ml x 1ml/1000µl = 0.0001 U/µl
2. Calculate amount of diluted insulin needed for dosage:
1.0mU/g x 1U/1000mU x 1µl/0.0001U = 10 µl/g
Eg. 20 g mouse:
10µl/g x 20 g mouse = 200 µl diluted insulin to inject.
10µl/g x body weight of mouse = dosage |
