Mouse Toe Identification Rationale
The use of mouse genetic models requires an efficient system of unique animal identifiers. The means that tissue must be obtained from each animal for genotyping and that each animal must be tagged so it can be quickly distinguished from other animals. A variety of methods have been employed to tag mice including ear tags, ear punches, tatoos and subcutateous chips. None of these approaches provide tissue so they must be coupled with a tissue biopsy, usualy a piece of tail tip, for genotyping. Alternatively, toe biopsy approach simultaneously provides tissue and a recognizable identifier to distinguish animals. Toe biopsies have other advantages, namely they do not cause chronic foreign body irritation, have less infection potential, can not fall off and require no specialized equipment.(1-6) The potential side effects of toe biopsies are similar to that of tail biopsies and are minimal if animals are biopsied in the recommended age range, and do not have health problems at the time of the procedure such as skin diseases, impaired immunity or thrombocytopenia. Like all live animal procedures, mouse identification requires proper training and prior IACUC approval. For genotyping procedures that are ammenable to a minimal amount of input material, cheek swabs are a less invasive alternative to toe or tail biopsies, but do need to be coupled with a permanent tagging procedure.(7) Approach
Mouse pups typically spend 3 weeks with their mothers before being weaned. During the first week they are too small for long term identifcation methods. Between 7 and 10 days of age their toes become well separated and can be biopsied with small dissecting scissors. At this age the tissue is still soft and the toe biopsy procedure appears to cause minimal distress. A 7 day old pup, for example, will typically object more to being picked up and separated from its littermates than it will to having its tail or toes biopsied. At older ages, mouse pups become more alert and the toe tissue becomes firmer from ossification. It is preferable to use younger animals because older animals require additional steps are taken to provide analgesia (e.g. with inhaled isoflourane) and ensure hemostasis. The use of anesthesitics significantly increases the risk of the procedure so biopsies should not routinely be performed on older animals. Procedure
1. The operator should wear gloves and protective clothing as per routine protocol.
2. Pick up a mouse pup by the scruff of the neck. When holding a mouse do not squeeze the animal so hard that its breathing is impaired. Do not hold it too loosely or it will squirm and become agitated. Taking care to hold a mouse properly is the most important aspect of ensuring its comfort and safety.
3. With a sharp pair of dissecting scissors remove a mouse toe and let the tissue fall into a 1.5 mL microcentrifuge tube. If more than one toe is being collected then pool them into the same tube. It is important dissect the tissue proximal to the interphalangeal joints so it can easily be recognized in the future.
4. Refer to the toe numbering scheme
to identify the animals with a unique ID. The system uses at most two toes from each foot and provides 0 - 9999 unique IDs. In practice smaller numbers can be used as long as care is taken not to add that two mice with the same toe IDs to the same cage. Cages cards and inventory records should include the complete unique identifier for each.
5. Place the pups which have been identified into a new cage to distinguish them from their littermates. When all of the pups have been identified, return the pups to their mothers cage.
6. Prepare DNA from the tissues according to the Tail DNA prep
7. Enter the new Pup IDs into the murine pathology database
(MPD) along with genotype information when it is available. References
1. Baron BW, Langan G, Huo D, Baron JM, Montag A. Squamous cell carcinomas of the skin at ear tag sites in aged FVB/N mice. Comp Med.
2. Cover CE, Keenan CM, Bettinger GE. Ear tag induced Staphylococcus infection in mice. Lab Anim.
3. Waalkes MP, Rehm S, Kasprzak KS, Issaq HJ. Inflammatory, proliferative, and neoplastic lesions at the site of metallic identification ear tags in Wistar [Crl:(WI)BR] rats. Cancer Res.
1987 May 1;47(9):2445-50.
4. Le Calvez S, Perron-Lepage MF, Burnett R. Subcutaneous microchip-associated tumours in B6C3F1 mice: a retrospective study to attempt to determine their histogenesis. Exp Toxicol Pathol.
5. Nadon NL, Draeger K. Genomic DNA analysis from mouse toe lysates. Transgenic Res.
6. 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.
7. Meldgaard M, Bollen PJ, Finsen B. Non-invasive method for sampling and extraction of mouse DNA for PCR. Lab Anim.