Table 2

Severity scale for ALS studies on transgenic mice with a mutant SOD1 gene

Severity	Description	Welfare issues during this stage
Level 1	Animals euthanised prior to disease onset, which is characterised by progressive weight loss or hind limb tremors.	No overt motor dysfunction. Phenotype is subclinical. Loss of motor function can be detected using rotarod or running wheels, but does not interfere with normal behaviour.
Level 2	Studies terminated at an early stage of disease: animals present trembling and weakness in hind limbs (by approximately 75 days) and mild body weight loss.	Minor. Loss of motor function can be detected using rotarod or running wheels, but has little interference with normal behaviour.
Level 3	Experiments terminated when animals are no longer able to reach food hopper or bottle spout. This occurs when animals reach a moderate (gait abnormalities and weakness) to severe (hind limb paralysis) stage of motor impairment (usually at 120–125 days).	Medium. Loss of motor function and body weight can be detected by monitoring (eg, by a clinical score sheet) and by checking self-righting ability. Refinement measures to address these welfare issues include provision of softer bedding material (eg, sawdust), elongated bottle spouts and mashed food on the cage floor.
Level 4	Animals euthanised after losing the ability to right themselves within 10–30 s after being laid on either side (one or both) or when percentage of weight loss reaches 15%–20% of peak body weight (usually at 130–140 days).	Major. Animals show severe locomotor impairment. Refinement as described for level 3.
Level 5	Animals are euthanised when reaching a moribund stage (complete paralysis) or allowed to die spontaneously.	Severe. At this stage, animals are unable to move, eat or drink. Animals which are not euthanised will die as a result of respiratory failure.

Each severity level exemplified from the most commonly used B6.Cg-TgN-(SOD1G93A) G1H mouse. Classification was based on the most severe endpoint used in each publication.
ALS, amyotrophic lateral sclerosis; SOD1, superoxide dismutase 1.