With the advancement of research and development of medical technology, there is an increase in the number of animals used in research. As millions of experimental animals are being used in different experiments worldwide, there is a lot of debate about the pain, distress, and death experienced by animals during scientific experiments. Experimental animals also require skilled manpower, time-consuming protocols and are costly. There are various alternatives to animal testing to overcome the drawbacks of animal experiments and avoid unethical procedures. Russell and Burch published principles of the Humane Experimental Technique which includes a strategy of 3 Rs (i.e. reduction, refinement, and replacement) in 1959. They introduced and defined the terms replacement, reduction, and refinement, which subsequently have become known as ‘alternatives’ or ‘alternative methods’ for minimizing the potential for animal pain and distress in biomedical research. Therefore, different methods and different alternative organisms are being used to implement the 3 Rs strategy. So, a brief account of these alternatives and the advantages associated is discussed in this review with examples. An integrated application of these approaches would give insight into the minimum use of animals in scientific experiments.