Date Adopted: May 8, 2018

Last Review Date: January 15, 2021

I.  Purpose

The purpose of this document is to provide guidelines to researchers regarding acceptable methods for marking animals to identify individuals (e.g., ear punching) and tissue collection for the purpose of rodent genotyping.

II.  Background

The Institutional Animal Care and Use Committee (IACUC) must approve all methods for tissue collection prior to performing procedures on animals. Toe punching, tail clipping, and toe clipping are acceptable methods of tissue collection for the purpose of genotyping mice and rats.

III.  Ear Tagging

Ear tagging involves the placement of a metal or plastic tag with a unique identification number or code to the base of the rodent’s ear.

A.  Animals must be 14 days or older.

B.  Ear tags should be disinfected prior to placement.

C. The site should be disinfected prior to placement with chlorhexidine or 70% 
     alcohol.

D. Ear tags should be placed in the lower half of the ear to facilitate normal ear 
     position and on the outer third of the ear to avoid the area with the highest 
     concentration of capillaries.

E.  Ear tags must be paced such that they do not cause a bend in the pinna, 
     interfere with the animal’s mobility, or be placed in such a manner that they can 
     catch on any part of the caging.

IV.  Ear Punching and Notching

This method involves punching a hole or making a notch in the ear pinna. Commercial ear punches are available and inexpensive. Ear notch or punch tissue remnants can usually provide enough tissue for genotyping.

A.  Animals must be14 days or older.

B.  Ear punching or notching does not require the use of anesthetics or analgesics, 
     however, the animal must be appropriately restrained to ensure proper technique.

C. The ear punch device must be disinfected between animals. These devices can 
     be autoclaved.

V.  Tail Clipping

This method involves amputating a very small segment of the distal tail. At < 21 days of age, the degree of ossification of the coccygeal vertebrate in the distal 5 mm is much less than that at 1 cm. After 21 days of age, the degree of ossification is similar along the entire length of the tail. The perception of pain is assumed to be more likely in bony vs. cartilaginous tissue. Therefore, tail clipping on mice or rats ≤ 21 days of age does not require anesthesia.

A.  Animals must be appropriately restrained during the procedure to minimize 
     trauma.

B.  Sterile sharp scissors (must be disinfected between uses) or a sterile blade per 
     animal can be used for the procedure.

C.  Only the distal 5 mm of the tail should be amputated.

D.  Hemostasis can be achieved by using a silver nitrate stick, Quick Stop powder, or 
     by applying a gauze sponge over the site with gentle pressure until bleeding stops

E.  Animals > 21 days of age or animals requiring a second tail sample must be 
     appropriately anesthetized using ketamine/xylazine, isoflurane, or a local 
     anesthetic during the procedure.

F.  Animals > 35 days of age that require tail clipping must be anesthetized 
     (ketamine/xylazine, isoflurane, or local anesthetic) and administered a systemic
      analgesic given at least once following the procedure. The AV may be consulted
     regarding the appropriate analgesics.

G.  If multiple tail clippings are required a maximum of 1 cm total tail length may be 
     amputated, with all tail clippings combined.

VI.  Toe Clipping

Toe clipping, as a method of identification of small rodents, should be used only when no other individual identification method is feasible and should be performed only altricial neonates and when combined with genotyping. The specific method(s) must be described in the Animal Use Protocol (AUP) and appropriate scientific justification provided. Under all circumstances, aseptic technique must be followed.

This method involves the removal of the distal phalangeal bone of one or more limbs. Toe clipping has the potential to induce pain and distress, and alter the animal’s gait and ability to feed.

A.  Only one toe per foot may be removed.

B.  Sterile sharp scissors can be used for this procedure (must be disinfected between 
     uses).

C. Hemostasis can be achieved using a silver nitrate stick, Quick Stop powder, or by 
     placing a gauze sponge over the site and applying gentle pressure until bleeding 
     has stopped.

D.  Toe clipping may only be performed in mice ≤ 12 days and rats ≤ 7 days old.

VII.  Other Methods of Identification

A.  Micro Chipping: A small microchip transponder is injected subcutaneously 
     between the scapulae of the rodent. The micro chip is detected by use of an 
     electronic reader.

B.  Micro-tattooing: A permanent mark using a needle and ink which is applied to the 
     tail, toes, or foot pads. The tattoo equipment must be disinfected between 
     animals.

C. Non-toxic markers: Sharpies can be used to mark the tail or fur of rodents, 
     however, the marks must be reapplied every 24 hours (or as needed) to ensure the 
     mark remains visible.

D.  Animal Marker™ is another product available which can be used on rodent fur. 
     The Animal Marker™ can remain visible for 6 -12 weeks.

VIII.  References

A.  Guide for the Care and Use of Laboratory Animals, 8^th Edition  pg. 75

B.  Hankenson FC Laire, Garzel LM, Fischer DD, Nolan B, and Hankenson KD. 
     Evaluation of tail biopsy collection in laboratory mice (Mus musculus): Vertebral 
     ossification, DNA quantity, and acute behavioral responses. J Am Assoc Lab Anim 
     Sci 47:10-18, 2008.

C. Vachon P. Anatomical and histological observations of fore- and hind limb toes in 
     adult mice after amputations performed at the age of two weeks. Can J Vet 
     Res 1998;62:311–13.