1.0 OBJECTIVE
1.1 To lay down a standard procedure to be followed for the humane and effective handling of laboratory animals to reduce pain and distress while giving euthanasia.
2.0 SCOPE
2.1 This Standard Operating Procedure (SOP) shall be applicable for giving euthanasia to laboratory animals in Animal House Facility.
3.0 RESPONSIBILITY
3.1 Study Director / In-charge- Animal House Facility
3.2 All personnel involved in giving anesthesia to laboratory animals
4.0 DEFINITIONS
4.1 Nil
5.0 PROCEDURE
5.1 General Considerations
5.1.1 Euthanasia
means an easy death and should be regarded as an act of humane sacrifice with minimum pain, fear, and distress.
5.1.2 Euthanasia
techniques must be consistent.
5.1.3 Animals should
not be sacrificed in the presence of other animals.
5.1.4 The method of euthanasia must be approved by the IAEC (Institutional Animal Ethics Committee).
5.2 Methods of Euthanasia
5.2.1 Carbon Dioxide Asphyxiation
5.2.1.1
General Considerations
5.2.1.1.1
The euthanasia chamber should allow complete visualization of the animals.
5.2.1.1.2 Do not
overcrowd the chamber; all animals in the chamber must be able to make normal postural
adjustments.
5.2.1.1.3
Compressed CO2
gas in cylinders is the only recommended source of carbon dioxide as it allows
the inflow of gas to the induction chamber to be controlled.
5.2.1.1.4
Pre-filling of
the chamber is not recommended as it has been shown that high concentrations of
CO2 is distressful to some animals.
5.2.1.1.5 Introduce 100%
carbon dioxide at the rate of 10-20% of the chamber volume per minute so as to
optimize reduction in distress.
5.2.1.1.6
After the
animals become unconscious, increase the flow rate to minimize the time to
death.
5.2.1.1.7 Unconsciousness
occurs within approximately 30 seconds, but animals should be left in the
chamber with the gas flowing for an additional 60 seconds. Neonates should be exposed
for approximately 5 minutes.
5.2.1.1.8
Death of the
animal(s) must be verified prior to experimental procedure or disposal.
5.2.1.1.9
A second
physical means of euthanasia: Cervical dislocation (rodents < 200 g) shall
be performed if required to assure death.
Note – Neonates of mice and rats are relatively
resistant to the effects of CO2 and other inhalants and may require
longer exposure times.
5.2.1.2
Procedure for operating Euthanasia setup for
Carbon Dioxide Asphyxiation
5.2.1.2.1
Pressure gauge
shall be checked for internal CO2.
5.2.1.2.2
If the main valve of the cylinder is closed, the valve shall be opened slowly by using the
appropriate cylinder key by rotating it anti-clockwise.
5.2.1.2.3
Absorbent
sheet/Cotton shall be kept in the euthanasia chamber before placing the
animals.
5.2.1.2.4
The animal
shall be placed in the euthanasia chamber.
5.2.1.2.5
Single-stage
regulator shall be turned on by rotating clockwise. Outlet pressure will be
displayed in the outlet pressure meter. The regulator shall be adjusted to maintain the
appropriate pressure based upon the required flow rate i.e 10 to 15-liter per
minute.
5.2.1.2.6
Flowmeter
valve shall be adjusted to release CO2 at the rate of 4.4 to 6.4 liters per
minute for the cylindrical euthanasia chamber and 2.8 to 4.0 liters per minute for the rectangular euthanasia chamber. The outlet valve shall be closed before placing
the animal in the euthanasia chamber for
euthanasia.
5.2.1.2.7
The animals
shall be exposed to CO2 for euthanasia and a single-stage regulator shall be
turned off by rotating anti-clockwise.
Note – Euthanasia chamber shall be cleaned
whenever found soiled and at the end of the procedure.
5.2.2 Chemical euthanasia
5.2.2.1.1 Anesthetics
shall be used in overdose for giving euthanasia to laboratory animals.
Regardless of the amount of chemical administered, the animal must be completely
non-responsive to noxious stimuli (hind paw pinch) before any physical means
are applied.
5.2.2.1.2
Barbiturate
anesthetics, phenobarbital, and ketamine injected intraperitoneal (IP) in
rodents produce rapid, smooth, and humane euthanasia.
5.2.3 Exsanguination
5.2.3.1 Exsanguination
shall only be carried out after the animal is completely non-responsive to
avoid the stress associated with extreme hypovolemia and the pain of incising
the deeper blood vessel.
5.2.3.2
Animals should
not be exsanguinated in the presence of other animals.
5.2.3.3
It shall be
performed if required during the necropsy procedure.
5.2.3.4
Abdominal the portion of an animal shall be cut with a V cut.
5.2.3.5
Vena cava shall
be identified and given an incision to drain out the blood.
5.2.3.6
Blood shall be
drained out in the waste collection container with running tap water to avoid
blood clotting.
5.2.4 Euthanasia by Cervical Dislocation
5.2.4.1
Animal shall
be removed from the cage and placed on the cage lid.
5.2.4.2
Procedure has
to take place in a quiet, clean environment, separate from other animals.
5.2.4.3 Restrain the mouse
by grasping the base of the tail with the thumb and forefinger, and by pressing the free thumb
and forefinger down firmly on the base of the head.
5.2.4.4 In one smooth
motion pull tail upward and outward quickly. The thumb and forefinger on the base of the head will roll down onto the neck.
5.2.4.5 Ensure that
there is a distinct dislocation of the vertebrae of the neck.
Note – Death must be established before disposal of
the carcass. This is assessed by lack of responses (pedal reflex, blink reflex) and loss of pulse.
5.2.5 The euthanasia procedure shall be documented as per current running version Format No. F…………………
6.0 ENCLOSURES
6.1 Formats
|
Sr. No. |
Format Title |
Format No. |
No. of pages |
|
1 |
Animal Euthanasia Record |
F…………………. |
01 |
7.1
SOP : Standard Operating Procedure
7.2
QAD : Quality Assurance Department
7.3
AHF : Animal House Facility
7.4
MBD : Microbiology
Department
7.5 SP : Study plan
8.0 REFERENCES
8.1
NIH Guide for Grants and Contracts. 7/17/2002, Notice: OD-02-062.
8.2 CCAC Guidelines on Euthanasia.
8.3 AVMA Panel on Euthanasia. 2000 Report of the AVMA Panel on Euthanasia. J Am Vet Med Assoc 2001, 218:669-696.
9.0
REVISION HISTORY
|
Sr. No. |
Change Control No. |
Reason for change |
|
|
|
|
END OF
DOCUMENT
You may like to read these links:
1. List of All SOPs and Documents for the Chemistry Laboratory
2. List of All SOPs and Documents for Quality Control
3. List of All SOPs and Documents For Quality Assurance Dept.
4. List of SOPs and Documents Required to Achieve NABL Accreditation
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