Insulin Tolerance Test: Difference between revisions

Line 5:

*Beaker for weighing mice

*Syringes

*0.1 U/mL humulin in PBS (make as 10uL of 100 U/mL in 10 mL, sterile filtered). This will correspond to 1 U/kg injections. This may need to be adjusted depending on the insulin sensitivity of the mice

*0.1 U/mL humulin in PBS (make as 10uL of 100 U/mL in 10 mL, sterile filtered). This will correspond to 1 U/kg injections. This may need to be adjusted depending on the insulin sensitivity of the mice. This is diluted from Humulin R-100 and is purchased through the veterinary staff.

*Timer

==Protocol==

*Remove food from mice for about 6h. Add do not feed tag to cages, or move cage to procedure room.

*Separate mice into individual cages and mark with mouse ID. Put a water bottle on the cage.

*Typically starve the mice at 9AM and aim to start injections at 3 PM

*Weigh mice, mark tails as necessary and take fasting glucose measurement via a tail clip.

*Weigh mice, mark tails if necessary and take fasting glucose measurement via a tail clip.

*Prepare glucose syringes with 10 uL per g mouse weight (ie for a 30g mouse, 300 uL).

*At approximately 1 min intervals, inject appropriate amount of ~~glucose~~ into interperitoneal cavity of the mouse.

*At approximately 1 min intervals, inject appropriate amount of insulin into interperitoneal cavity of the mouse.

*Immobilize mouse and restrain tail with one hand

**Immobilize mouse and restrain tail with one hand

*Aim needle between the midline and the hip bone

**Aim needle between the midline and the hip bone

*Insert syringe (do not inject) into cavity

**Insert syringe (do not inject) into cavity

*~~Pull up syringe and ensure that only air is aspirated. If urine or blood appears in the syringe, start over~~

**Eject syringe.

*Eject syringe.

*At desired intervals (normally 15, 30, 45, 60, 75, 90, 105 and 120 min), take blood glucose measurements from tail vein. If needed re-snip the tail vein. When measuring glucose just lift the tail of the mouse, while leaving it in the cage, rather than removing and restraining the mouse.

*At desired intervals (normally 15, 30, 45, 60, 75, 90 and 120 min), take blood glucose measurements from tail vein

*Analyse data by both % change from fasting glucose and absolute values

@@ Line 5: / Line 5: @@
 *Beaker for weighing mice
 *Syringes
-*0.1 U/mL humulin in PBS (make as 10uL of 100 U/mL in 10 mL, sterile filtered). This will correspond to 1 U/kg injections.  This may need to be adjusted depending on the insulin sensitivity of the mice
+*0.1 U/mL humulin in PBS (make as 10uL of 100 U/mL in 10 mL, sterile filtered). This will correspond to 1 U/kg injections.  This may need to be adjusted depending on the insulin sensitivity of the mice.  This is diluted from Humulin R-100 and is purchased through the veterinary staff.
 *Timer
 ==Protocol==
 *Remove food from mice for about 6h. Add do not feed tag to cages, or move cage to procedure room.
-*Separate mice into individual cages and mark with mouse ID.  Put a water bottle on the cage.
 *Typically starve the mice at 9AM and aim to start injections at 3 PM
-*Weigh mice, mark tails as necessary and take fasting glucose measurement via a tail clip.
+*Weigh mice, mark tails if necessary and take fasting glucose measurement via a tail clip.
 *Prepare glucose syringes with 10 uL per g mouse weight (ie for a 30g mouse, 300 uL).
-*At approximately 1 min intervals, inject appropriate amount of glucose into interperitoneal cavity of the mouse.
+*At approximately 1 min intervals, inject appropriate amount of insulin into interperitoneal cavity of the mouse.
-*Immobilize mouse and restrain tail with one hand
+**Immobilize mouse and restrain tail with one hand
-*Aim needle between the midline and the hip bone
+**Aim needle between the midline and the hip bone
-*Insert syringe (do not inject) into cavity
+**Insert syringe (do not inject) into cavity
-*Pull up syringe and ensure that only air is aspirated. If urine or blood appears in the syringe, start over
+**Eject syringe.
-*Eject syringe.
+*At desired intervals (normally 15, 30, 45, 60, 75, 90, 105 and 120 min), take blood glucose measurements from tail vein.  If needed re-snip the tail vein.   When measuring glucose just lift the tail of the mouse, while leaving it in the cage, rather than removing and restraining the mouse.
-*At desired intervals (normally 15, 30, 45, 60, 75, 90 and 120 min), take blood glucose measurements from tail vein
 *Analyse data by both % change from fasting glucose and absolute values