3Rs advice for project licence applicants: refinement

Refinement refers to methods which minimise animal suffering and improve welfare. It is the responsibility of the project licence holder to ensure that any regulated procedures specified in the licence cause the least pain, suffering, distress or lasting harm to the animals and are the most likely to provide satisfactory results. Information about refinement is mostly given in the “Refinement and Protocols” sections of the licence application.

Sometimes applicants misunderstand the guidance text provided for the Refinement section and repeat information given in the Replacement section, focusing on why non-animal alternatives are not suitable. Instead, you should explain why you have chosen this animal species, model and method to answer your scientific question and why this approach is the most refined for the animals in comparison to other animal-based methods you could have used to address your scientific question.

Applicants may also misunderstand the definition of refinement, confusing it with refinements to optimise their experimental design/data output. Refinements in the context of the project licence application should always refer to those which will benefit the animals and improve their welfare.

Systematic reviews

Systematic reviews can be used to select the most appropriate animal model and determine if less severe procedures are as predictive as more severe alternatives. (They can also be used to inform effect sizes and variability used in sample size calculations). Our webinar on ‘Systematic reviews of animal studies’ provides an introduction to systematic reviews, including the type of research questions that can be addressed, the steps included, tools available and the 3Rs benefits of systematic reviews. One such tool, the Systematic Review Facility (SyRF) is a free-to-use platform funded by the NC3Rs which supports researchers to carry out systematic reviews and meta-analysis of animal studies. It should be noted that systematic reviews can be time-consuming to conduct and typically need to be funded, however they can be enormously useful as part of a wider programme of research. Preparing a project licence (or grant application) provides an opportunity to consider whether a systematic review of the literature would be helpful and what research questions could usefully be addressed using this approach.

General refinement advice

When outlining the refinements to be used, applicants should not forget to include smaller, incremental refinements which impact on animal welfare. These could be refinements to the procedure itself (e.g. choosing the most refined route of administration; habituation to procedures; acclimatising animals to restraint tubes in the home cages) or applying husbandry-based refinements (e.g. providing food on the cage floor for mouse-models of arthritis; using non-aversive mouse handling methods).

When suggesting refinements, ensure these have been discussed with the Named Persons, such as the Named Animal Care and Welfare Officer (NACWO) and Named Veterinary Surgeon (NVS), and that they are both relevant to the model and feasible within your facility. Further guidance on planning for experimental procedures, is given in NORECOPA’s PREPARE guidelines.

Humane endpoints

A key component of refinement is the implementation of humane endpoints. We have a humane endpoints webpage that provides information on how to establish appropriate humane endpoints that minimise animal suffering. Further advice is given in the Humane Endpoints website of the 3Rs Centre at Utrecht University.

It is important to remember that in many studies pain and distress can be avoided by the identification of non-clinical scientific endpoints that occur before any observable suffering or clinical signs of a condition. Examples of this might include the use of imaging technologies to assess internal tumour burden or the use of study-specific biomarkers in serum. By identifying earlier scientific endpoints this will allow more refined humane endpoints to be implemented, thereby minimising pain and suffering in the models used.

Welfare assessment protocols and score sheets can be useful tools to monitor adverse effects and determine when humane endpoints have been reached. The experience of the Named Persons and animal technicians can be invaluable when creating welfare score sheets that accurately reflect your model by recording relevant adverse effects and incorporating the earliest humane endpoints. The score sheet/s need not be included in the licence application itself but should be shared with the AWERB.

Disease-specific guidance

The NC3Rs has led a series of expert working groups which focus on specific disease models that have the potential to cause severe suffering and opportunities for their refinement. These include:

Use the ‘Search our science’ box in the What we do area of our website to identify opportunities to refine the use of other disease models and procedures.

The RSPCA has also produced a series of reports on refining severe models and procedures.

The Workman et al. (2010) guidelines for the welfare and use of animals in cancer research are an invaluable resource, setting out appropriate humane endpoints and relevant clinical signs to monitor.

Although the examples above are disease-specific guidelines, certain refinements can have wider applications to other models (e.g. social housing post-surgery; wet mash to encourage eating; extra monitoring of animals with co-morbidities).

Refinement resources

The NC3Rs website provides a number of resources to help with further refinement of your research. These include advice and guidance on common procedures such as blood sampling and microsampling, and the evidence base to support the single use of needles as a refinement. We also have species specific pages dedicated to non-aversive mouse handling and genetically altered mice.

Our resources hub also contains e-learning resources on conducting welfare assessments and on euthanasia and anaesthesia. Training material on the Procedures with Care resource focuses on the administration of substances to mice and rats and aseptic technique in rodent surgery.

We also have dedicated resources focused on applying the 3Rs in toxicology and regulatory science, with one of the most useful documents being the LASA and NC3Rs guidance on dose level selection for regulatory general toxicology studies for pharmaceuticals.

Page last updated: 6 July 2020