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What are the 3Rs?

The 3Rs are a widely accepted ethical framework for conducting scientific experiments using animals humanely:

  • Replacement - use of non-animal methods

  • Reduction - methods which reduce the number of animals used

  • Refinement - methods which improve animal welfare

The 3Rs principles were first introduced in Russell and Burch's 1959 book 'The principles of humane experimental technique'.

Species selection has a bearing on each of the 3Rs. The NC3Rs has adopted the following 3Rs definitions:


Replacement refers to methods that avoid or replace the use of animals defined as 'protected' under the Animals (Scientific Procedures) Act 1986, amended 2012 (ASPA) in an area where they would otherwise have been used.

'Protected' animals are all living vertebrates (other than man), including some immature forms, and cephalopods (e.g. octopus, squid, cuttlefish).

Replacement methods can be:

  • Absolute replacements - those which do not involve animals at any point
  • Relative replacements - those which avoid or replace the use of 'protected' animals

Examples of replacement methods include:

  • Computer modelling
  • Human volunteers, e.g. for non-invasive imaging studies 
  • In vitro methodologies, utilising:

- established human or animal cell lines
- animal cells, tissues and organs from animals killed by a humane method (see Schedule 1 of the ASPA)
- abattoir material from the meat industry

  • Invertebrates, such as Drosophila (fruit fly) and nematode worms
  • Immature forms of vertebrates:

- mammal, bird and reptile embryos, up to the last third of their gestation or incubation period 
- larval forms of amphibians and fish, until the stage where they become capable of independent feeding
- cephalopods until the point at which they hatch


Reduction refers to methods which minimise animal use and enable researchers to obtain comparable levels of information from fewer animals or to obtain more information from the same number of animals, thereby reducing future use of animals.

Examples of reduction methods include: 

  • Improved experimental design and statistical analysis
  • Modern imaging techniques
  • Sharing data and resourcesFig 3








Non-invasive, whole body imaging of small animals using techniques such as X-ray, CT, SPECT, PET and MRI, is helping to reduce the number of animals used in basic research and drug development. The same animal can be imaged multiple times in order to monitor visually, often in real time, the progression or regression of infection or disease. This avoids the need to sequentially sacrifice animals at different time points, allowing significant reductions in the number of animals used per study. The image above shows biophotonic imaging (BPI) of a single mouse infected inter-peritoneally with a bioluminescently engineered strain of the bacterium, Pseudomonas aeroginosa.


Refinement refers to improvements to scientific procedures and husbandry which minimise actual or potential pain, suffering, distress or lasting harm and/or improve animal welfare in situations where the use of animals is unavoidable.

Refinement applies to the lifetime experience of the animal. There is evidence that refinement not only benefits animals, but can also improve the quality of research findings.

Examples of refinement methods include:

  • Non-invasive techniques
  • Appropriate anaesthetic and analgesic regimes for pain relief
  • Training animals to voluntarily co-operate with procedures (e.g. blood sampling) so that they have greater control over the procedure and are less stressed 
  • Accommodation and environmental enrichment which meets the animals' physical and behavioural needs (e.g. providing opportunities for nesting for rodents)

Red mouse house

The 'mouse house' is a refinement developed at the MRC National Institute for Medical Research. The transparent, red, plastic house enables the mice to perform natural behaviours such as nesting, hiding and climbing, which is important for their welfare. The house appears dark to the mice, yet the transparent walls mean that animal care staff can see the mice at all times and so are able to carry out their daily checks without disturbing them.

Further information on these and other 3Rs methods is given in our Information Portal, Invited Articles and Research Review.

Related Documents

link arrow Finding alternatives: an overview of the 3Rs and the use of animals in research (PDF, 24KB)
link arrow The challenge of animal research (PDF, 681KB)

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