Alphavirus, a genus of mosquito-borne arboviruses, cause endemic disease globally and are of serious public health concern. Arthritogenic alphaviruses such as Mayaro virus, Chikungunya virus, O'nyong nyong virus and Ross River virus are prevalent mainly in Africa and Asia and are associated with debilitating arthritic disease. Whereas encephalitic alphaviruses such as Western, Eastern, and Venezuelan equine encephalitis viruses are mainly found in the Americas and lead to human neurological disease and fatal encephalitis. A range of the host’s innate and adaptive immune responses are shown to be activated during alphavirus infection, including the induction of cellular responses associated with B cells, T cells, macrophages, Natural Killer cells as well as the induction of cytokines, chemokines, and complements. Significant progress in alphavirus research has been achieved in recent years as novel advanced experiment methods and techniques have been applied in this field, such as poly-protein pulse-labelling, genome-wide CRISPR screening, reverse genetics system, and genetic modification for vaccine development among others. Hence, summarizing and sharing these validated methodologies is helpful not only to future research in alphavirus but could also shed light on investigations of other viruses and associated diseases. We hope this practical visualized Methods Collection could promote basic research in virology and antiviral immunity, encourage worldwide research collaborations, as well as aid the development of novel approaches to counteracting infectious diseases.