Inadequate exposure time to CO2 may result in animals that appear dead but can awaken from deep anesthesia. One of the following procedures MUST be followed to assure death. This should match the description in your IACUC approved ACUP.

  • Bilateral pneumothorax of deeply anesthetized or dead animals—
    • Open the chest wall/thorax
    • Open the abdomen and incise the diaphragm
  • Cervical dislocation – animals that are non-responsive to stimulus may be killed by cervical dislocation. This procedure is limited to mice, or rats less than 200g and must be described in the approved ACUP
  • Decapitation-as described in the approved ACUP
  • Experimental procedures on deeply anesthetized or dead animals which assure death by another means (e.g. exsanguinations, vital organ removal)


  • 2007 AVMA ”Guidelines on Euthanasia”
  • Report of the ACLAM Task Force on Rodent Euthanasia, American College of Laboratory Animal Medicine, 2005
  • Guide for the Care and Use of Laboratory Animals, 2011

IACUC approved May 2011