Identification methods in newborn C57BL/6 mice: a developmental and behavioural evaluation

Abstract

The use of group-housed rodents in many fields of biomedical research imposes a need to identify individuals in a cage. Few studies have been designed to assess possible negative effects of identification methods of newborn mice on their development and wellbeing. In the present study, three different identification methods were applied to newborn C57BL/6J mice on postnatal day ( pnd) 5 (toe clipping, toe tattoo ink puncture and subcutaneous implantation of a small transponder). All identification methods used proved to be effective for long-term marking of individual animals. Newborn mice showed the least reaction to toe clipping followed by toe tattoo ink puncture and transponder implantation was the most distressful individual identification procedure in newborn mice. Importantly, clipped toe tissue proved to be enough for genotyping purposes. No overall consistent differences in somatic and neurological reflex development during the postnatal period were shown as a result of the newborn individual identification procedures used. Further, none of the methods interfered significantly with the adult animals' general normal behaviour (e.g. ability to move, grasp, climb) and sensory -motor functions as assessed with a simplified SHIRPA battery of tests, as well as Rotarod and Elevated Plus Maze tests. Postmortem thymus and adrenal gland weights gave no indication of chronic stress as a consequence of the identification method. We conclude that toe clipping might even be advisable in newborn mice at a very young age, when genotyping is needed. Toe tattoo ink puncture is also a good identification method for newborn mice and transponder implantation should only be used in older newborns or applied at weaning.

Report of the Federation of European Laboratory Animal Science Associations Working Group on animal identification

“…20 None of the methods exhibited any postnatal effects. The authors recommended the use of toe clipping in young pups and microchips only after weaning.…”

“…It is likely that too much of the toe was cut on day 3 due to the small size. 26 No effects on climbing abilities at weaning, 11 or as adults, 20 were found in mice with distal phalanx removal.…”

FELASA guidelines for the refinement of methods for genotyping genetically-modified rodents

“…Distal phalanx biopsy consists of the removal of the distal phalanx of a newborn animal, that is then used as a source of DNA. This method also provides a means of identification, and is described in detail in the literature [11][12][13] and in the report of the FELASA Identification Working Group.…”

“…12,13 Conversely, tail biopsy has been shown to affect mice, at least in the short term. 10,15 Although the technique is possible earlier, postnatal day (PND) 7 seems to be the preferred age for distal phalanx biopsy.…”

Rapid Genotyping of Animals Followed by Establishing Primary Cultures of Brain Neurons

“…Tattooing newborn animals is advantageous because it can be carried out rapidly and lasts for much of the animal's life 7,[22][23][24][25] . Tattooing on the paw pads can be better than toe clipping 23 with respect to preserving testable behaviors, for example in suspension or gripping 22,26 , although some studies have not noted such deficiencies (e.g., 25 ). There were no instances of mothers rejecting or cannibalizing pups after tattooing.…”