The euthanasia is one of the most studied, debated and controversial topics in the laboratory animal science area. All over the world, the ideal way to perform euthanasia in laboratory animals is sought, seeking, in summary, three main points, animal welfare during the procedure; minimal influence on the animals biological system, not to interfere with test results and accessibility and low cost. In Brazil, after the validity of Law No 11.794-2008, that regulated the use of animals for didactic and scientific purposes, structuring the Animal Experimentation Control Council (CONCEA). In its resolutions, CONCEA standardized the application of accepted euthanasia methods, mainly overdose of injectable anesthetics and restricted use of carbon dioxide (CO2 ). The aim of our study was to evaluate within the rules of CONCEA and using non-invasive and invasive parameters which would be the best euthanasia promoter in adult Swiss Webster male mice. We compared the use of CO2 (suggested a substitution in the in sufflation valve) Isoflurane saturation, a combination of induction in an environment saturated with isoflurane and progressive CO2 in sufflation and application of the association Ketamin and Xilazin, in overdose. Our results, clearly, demonstrate that the most humane method of inducing the death of the animal is that which is the fastest (Speed Death); does not promote pain, discomfort or distress in the animal; minimally influences the biological systems of the animals and has a low acquisition cost. From our results it was possible to suggest that the combination of isoflurane (5%) and CO2 (1 L/min) is the most efficient methodology for performing euthanasia in mice. Because it is extremely fast, mice does not show discomfort, does not influence the biological systems of the animal after death (except the Central Nervous System) and has a low cost with relative ease of access. Thus, we believe that with this study, we can demonstrate that through projects, studies and tests we can suggest new and reliable methodologies for the application of the welfare of laboratory animals.