Today, the idea of replacement is more restrictive and usually refers to the use of either tissue culture or some other experimental system that does not require either disturbing or killing an animal. Even in these cases, however, their is some level of disturbance to animals. The question of what constitutes an animal in the alternatives framework is exemplified by attitudes toward Limulus, an invertebrate. Generally, invertebrates are perceived to be replacements. However, a few papers have discussed the sentience of insects and, recently, the British Home Office decided to add the octopus to the category of protected research animals.
Reduction refers to modifications in procedures that allow fewer animals to be used.
These include the Fixed Dose Procedure, the Acute Toxic Class Method, and the Up and Down Procedure, all of which typically use fewer than 20 animals per test. Some observers would consider these cases to be examples of replacement techniques, while others would consider them to be examples of reduction in numbers. Refinement refers to efforts to decrease the incidence or severity of painful or stressful procedures for animals which still are used in specific tests or research. An alternative is a new research technique that uses no or fewer animals or causes less animal distress. One of the features of biomedical and biological science is the increasing sophistication of research technology and the expanding ability to answer ever more complicated and detailed questions.