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Insulin tolerance test (ITT)

This protocol may be used to perform ITT on mice.

Category
Protocol
Record History
Added on June 11, 2010 at 10:02 AM by Lindner, Jill
Modified on July 1, 2010 at 3:21 PM by Lindner, Jill
Shared with (contributions)
Public: Shared Collection

Reagents, supplies, and equipment:

  • #21 surgical blades
  • 21 ½ G needle
  • insulin syringe
  • 5 ml syringe 
  • human insulin 100 IU
  • 0.9 % sterile saline
  • BD logistic glucometer
  • BD glucose test strips
  • Clean mouse cages (with normal bedding: dried corn)
  • bench paper
  • 250 ml plastic beaker
  • 70% EtOH

Procedure: Day 1

  1. Bring mice to lab around 5:30 PM
  2. Disinfect work area with 10% Clorox, then 70% EtOH
  3. Prepare work area with balance, clean bench paper, needles, blades, glucometer, and insulin syringes.
  4. Allow mice to rest in the lab O/N for ~ 15 hr.  Provide food and water as normal.

Procedure: Day 2

  1. Fast mice for 6 hr between 9:00 AM - 3:00 PM.  (Place mice in a clean cage without food and with H2O).
  2. Weigh  mice at 2:30 PM.
  3. Calculate the insulin dose for each mouse (based on body weight).
  4. Prepare 0.25 IU insulin solution by diluting insulin in sterile saline (and sterile tubes) @ 1:400; mix by vortexing..
  5. Dispense the required volume of insulin solution for each mouse into separate 1.5 ml tubes (Volume is calculated as follows: 0.75 IU insulin/g BW).  Vol (μl) = 3 x BW
  6. Measure blood glucose of the 1st mouse from tail blood by cutting < 5 mm of the tail tip.
  7. Perform intraperitoneal injection (i.p.) of insulin.  This is time 0' (3:00 PM).  Record blood glucose level(s).
  8. Continue i.p. injections on subsequent mice, maintaining at least a 1 min interval between animals.
  9. Repeat blood glucose measurement at 15', 30', 45', 60', and 90' by reopening the tail wound each time. There is no need to re-cut the tail.
  10. Upon completion, provide food to each mouse and return the mice to the mouse colony.

Written by: Yanyun Gu 5/10

 

Attachment

Document itt_worksheet.doc - Added on June 11, 2010 at 11:12 AM by Jill Lindner