Issuing Office(s)
_____IACUC____________
Responsible University Officer(s)
______UNC IACUC ______
UNIVERSITY STANDARD
Title
UNIVERSITY OF NORTH CAROLINA AT CHAPEL HILL
STANDARD FOR RODENT BLOOD WITHDRAWAL AND TAIL BIOPSY
Introduction
PURPOSE
The standards and procedures described below provide guidance to all researchers and animal
handlers for performing blood withdrawals and tail biopsies in mice and rats.
SCOPE OF APPLICABILITY
All personnel engaged in the hands on collection of blood and/or tail tissue from rats and mice.
The UNC-CH IACUC expects that anyone involved in animal work at the University will
comply with this Standard and the University Policy on the Care and Use of Vertebrate
Animals for Research, Training and Teaching Purposes. Requests for exceptions to this
Standard must be reviewed and approved by the IACUC and/or DCM Management.
TRAINING
Your Laboratory Animal Coordinator (LAC) if qualified, may be able to train others in this
technique, or you can contact the IACUC at 919-966-5569 (iacuc@med.unc.edu) to arrange
for training.
See the IACUC Mouse Handling & Techniques and Rat Handling & Techniques packets for
more detailed instructions and diagrams.
Standard
Acute or Single Blood Withdrawal: The maximum survival amount of an acute blood
withdrawal is 1% of the lean body weight. [e.g.; For a 20 gram adult mouse, no more
than 200 µl (4 X 50 µl micro capillary tubes), may be withdrawn]
Chronic Blood Withdrawal: For sequential blood sampling (over a period of time), the
maximum aggregate survival blood withdrawal for most mammals is 1.5% of lean body
weight over a 14-day period. The above guideline for acute or single blood withdrawal
still holds at any given time point within this 14-day period.
To facilitate blood collection, warm the rodent first. When using the tail veins or artery,
you may dip the tail in warm water (45°C). The entire animal can be warmed with a
carefully placed heat lamp for 5-10 minutes or by placing the housing cage on a
Page 1 of 5
Policy Title: UNIVERSITY OF NORTH CAROLINA AT CHAPEL HILL STANDARD ON …
Effective Date: July 26, 2002
Last Revised: 9/19/03, 7/27/04, 10/21/05, 6/14/06, 3/13/09, 02/18/13, 04/07/17, 12/2017
Issuing Office(s)
_____IACUC____________
Responsible University Officer(s)
______UNC IACUC ______
circulating water pad. Alternatively, alcohol may be used initially as a vasodilator, but it
should not be used on broken skin.
Table 1: Approximate Blood Sample Volumes for a Range of Body Weights
1% CBV (l) 7.5% CBV (l) 10% CBV (l)
Body weight (g) *CBV(l)
every 24 hrs† every 7 days† every 2 - 4wks†
20 1100 - 1400 11- 14 82 - 105 110 - 140
25 1370 - 1750 14 - 18 100 – 130 140 -180
*Circulating blood volume †Maximum sample volume for that sampling
** adapted from NIH Guidelines for Survival Bleeding offrequency
Mice and Rats
Methods:
1. Submandibular:
A relatively simple way to obtain blood (usually only used to collect blood from mice) is to puncture
the area behind the hinges of the jawbones. The superficial temporal vein is a large vessel
positioned behind the eye, which can be traced backward to the temporal vein, the maxillary vein,
and finally, the jugular vein.
Scruff the mouse and pierce the skin in the relevant area. A mouse bleeding lancet is strongly
recommended for use. However, an 18 gauge needle may also be used.
2. Saphenous Vein:
This method of obtaining blood is often used when a series of small samples is required.
Restrain the animal and shave the caudal surface of the thigh. The saphenous vein can be seen
in this area. It is advantageous to apply a lubricant to prevent wicking. Place a tourniquet above
the knee and enter the vein with a needle. This method of blood withdrawal does not require
anesthesia, however, the method of restraint is cumbersome.
3. Tail Artery / Vein (Tail Nick Bleed):
Tail veins and artery can be used for serial bleedings. Use the central tail artery or lateral tail
veins. Anesthesia is not required for tail vein nick bleed or tail artery bleed.
Tail Nick Bleed: When bleeding from either of the lateral tail veins, a 22-gauge needle or lancet
is inserted into the vein, midway up the tail. You may collect blood with micro capillary tubes, or
a micropipette.
Tail Artery Bleed: Most common in rats. Blood may be withdrawn from the ventral tail artery
using a 22-gauge (or smaller) needle; let the blood drip into the collection vessel. A tourniquet
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Policy Title: UNIVERSITY OF NORTH CAROLINA AT CHAPEL HILL STANDARD ON …
Effective Date: July 26, 2002
Last Revised: 9/19/03, 7/27/04, 10/21/05, 6/14/06, 3/13/09, 02/18/13, 04/07/17, 12/2017
Issuing Office(s)
_____IACUC____________
Responsible University Officer(s)
______UNC IACUC ______
placed at the base of the tail will facilitate bleeding. Anesthesia is not required but is highly
recommended for ease of collection.
4. Tail Clip Bleed and/or Tail Biopsy for Genotyping:
Performed on unanesthetized or anesthetized animals depending on amount of tissue needed
(see below):
Anesthesia is optional for the removal of up to 4mm from the tail tip. It is strongly
recommended that no more than 2mm be removed at a time. Anesthesia may be used
as a means of animal restraint and its use must be described in the approved protocol.
Depending on the age of the animal, removal of greater than 4mm from the tail tip may
involve cutting into the vertebral column. Therefore, anesthesia is always required when
removing this much tail, irrespective of the age of the animal. The use of anesthesia must
be described in the approved animal care application.
Requests to perform tail biopsies or successive tail cuts totaling greater than 4mm
without anesthesia must be scientifically justified and must receive IACUC approval
prior to implementation.
The IACUC has approved the tail cut method for both rats and mice to obtain blood and/or tissue.
This method must be described in the animal use application and approved by the IACUC prior
to use. See procedure below.
1. Place animal in approved animal restrainer. (Experienced handlers may be able to
perform technique in habituated animals with light or no restraint).
2. Remove any bedding material or feces from the tail.
3. Place the animal on a clean work surface.
4. Using a fresh scalpel blade, cut 1-2 mm of the distal tail at an angle perpendicular to
the work surface.
5. Apply gentle pressure proximal to the collection site to occlude venous return and ease
collection.
6. Apply gentle digital pressure to the wound for 30-45 seconds with a clean gauze pad
to stop bleeding, before placing animal back in its home cage. For persistent bleeding,
apply a silver nitrate stick, styptic powder or a cautery pen to the wound to stop
bleeding.
7. Serial blood samples can be obtained over a short time frame by gently removing the
scab without performing an additional cut.
8. Only the fleshy portion of the tail tip should be cut. Cutting into the vertebrae is NOT
permitted. As only a small portion of the tail does not contain vertebrae, the use of the
tail cut procedure should be limited.
5. Retro Orbital Bleeding:
Retro-orbital or orbital sinus/plexus bleeding is permitted in rats, mice, gerbils, guinea pigs,
hamsters. The IACUC will permit orbital sinus bleeding when it is scientifically justified, performed
with appropriate technique and anesthesia. Veterinary staff experience indicates that this method
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Policy Title: UNIVERSITY OF NORTH CAROLINA AT CHAPEL HILL STANDARD ON …
Effective Date: July 26, 2002
Last Revised: 9/19/03, 7/27/04, 10/21/05, 6/14/06, 3/13/09, 02/18/13, 04/07/17, 12/2017
Issuing Office(s)
_____IACUC____________
Responsible University Officer(s)
______UNC IACUC ______
may lead to orbital damage, blindness and potentially death if not performed correctly. The IACUC
encourages the primary use of the submandibular, tail artery or veins; specifically the nick or cut
techniques. These methods are less likely to harm the animal and may be used repeatedly for
bleeding.
LACs may not train in this technique so training and certification must be obtained from IACUC
Training and Compliance team or DCM veterinary services.
Alternating eyes for each bleeding is mandatory, and a week must separate each bleeding
(unless the final bleed is performed as a terminal procedure – animal does not recover
from anesthesia). A maximum of two (2) bleedings per eye is permitted. Maximum volume
withdrawn within a two week period is 1.5% body weight. Orbital sinus bleeding requires
training and must be performed on anesthetized animals only with IACUC approval.
“Following blood collection, the eyelids should be held closed for a few seconds to allow the
punctured blood vessel to clot. It is also common practice to place a small amount of ophthalmic
ointment into the eye following this procedure.” excerpt from Laboratory Animal Technician
Training Manual.
6. Cardiac Puncture: Always a terminal procedure conducted under anesthesia or
immediately following euthanasia!
Cardiac puncture as a method of blood withdrawal permitted in all species provided the following
conditions are met:
1. Animal is under a surgical plane of anesthesia or deceased when procedure is
conducted.
2. Animal is NOT allowed to recover from anesthesia following the puncture.
3. If the animal is euthanized prior cardiac puncture, training and certification in the
technique is not required.
4. A needle is inserted into the heart and blood is extracted until a sufficient volume is
collected or the animal is exsanguinated. This procedure must be followed by a
physical euthanasia method.
EXCEPTIONS
Requests for exceptions to this Standard must be reviewed and approved by the IACUC and/or
DCM Management.
Definitions
IACUC: Institutional Animal Care and Use Committee
DCM: Division of Laboratory Animal Medicine
OACU: Office of Animal Care and Use
University Standard: The minimum acceptable limits or rules used to achieve Policy
implementation, enforceable by the IACUC.
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Policy Title: UNIVERSITY OF NORTH CAROLINA AT CHAPEL HILL STANDARD ON …
Effective Date: July 26, 2002
Last Revised: 9/19/03, 7/27/04, 10/21/05, 6/14/06, 3/13/09, 02/18/13, 04/07/17, 12/2017
Issuing Office(s)
_____IACUC____________
Responsible University Officer(s)
______UNC IACUC ______
Blood Withdrawal: Removal of blood, usually by venipuncture
Related Requirements
EXTERNAL REGULATIONS AND CONSEQUENCES
UNIVERSITY POLICIES, STANDARDS, AND PROCEDURES
For more detailed guidance, please refer to the University Policy on the Care and Use of
Vertebrate Animals for Research, Training and Teaching Purposes.
Contact Information
Subject Contact Telephone Email
IACUC Protocol IACUC 919-966-5569 iacuc@med.unc.edu
and Training
Veterinary Services DCM 919-966-2609
Important Dates
Effective Date and title of Approver: 07/26/2002; UNC IACUC
Revision and Review Dates, Change notes, title of Reviewer or Approver: 01/16/03,
07/24/04,10/21/05,06/14/06/03/13/09,02/08/13, 12/1/2017; UNC IACUC
Approved by: UNC IACUC
Dr. Mitchell Picker 12/2017
UNC IACUC Chair
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Policy Title: UNIVERSITY OF NORTH CAROLINA AT CHAPEL HILL STANDARD ON …
Effective Date: July 26, 2002
Last Revised: 9/19/03, 7/27/04, 10/21/05, 6/14/06, 3/13/09, 02/18/13, 04/07/17, 12/2017
