The 3Rs – Reduction, Replacement and Refinement – are embedded into the legislation and guidelines governing ethical rigour of animal use in experiments. Here, we consider the advantages of adopting key aspects of the 3Rs into experimental biology, represented mainly by the fields of animal behaviour, neurobiology, physiology, toxicology and biomechanics. Replacing protected animals with less sentient forms or species, cells, tissues or computer modelling approaches has been broadly successful. However, many studies investigate specific models that exhibit a particular adaptation, or a species that is a target for conservation, such that their replacement is inappropriate. Regardless of the species used, refining experiments by ensuring the health and wellbeing of animals prior to and during experiments is crucial for the integrity of the results and legitimacy of the science. Although the concepts of health and welfare are developed for model organisms relatively little is known regarding non-traditional species that may be more ecologically relevant. Studies should reduce the number of animals in experiments by employing the minimum suitable sample size. Presently this is often calculated using power analyses, which is associated with making statistical inferences based on the P value, yet P values often leave scientists on shaky ground. We endorse focussing on effect sizes accompanied by confidence intervals as a more appropriate means of interpreting data; in turn, sample size could be calculated based on effect size precision. Ultimately, the appropriate employment of 3Rs principles in experimental biology empowers scientists in justifying their research, and results in higher quality science.