This thesis aimed to provide evidence-based recommendations for improving cat welfare during handling. First, a cat handling questionnaire suggests that veterinarians (n=344) and non-veterinarian staff (n=944), commonly use restraint involving immobilization for examinations and procedures when handling fearful and aggressive cats. Participants were less likely to use full-body with scruff restraint (known negative) on fearful (F) and aggressive (A) cats if they work: at a Cat-Friendly Practice (F: P=0.0001; A: P=0.0001), in Canada (F: P<0.0001; A: ns), as a veterinarian (F: P<0.0001; A: ns), or as a non-veterinarian who graduated in 2005 or earlier (F: P<0.0001; A: P<0.0001). Given a lack of scientific evidence to inform good handling, methodologies were validated to assess cat responses to handling by comparing full-body (known negative) and passive (control) restraint. Cats were categorized as friendly or unfriendly based on interactions with a stranger, given the hypothesis that unfriendly cats would display more avoidance behaviours than friendly cats. Conditioned place aversion (CPA) was validated for use with laboratory (n=10) and friendly shelter (n=26) cats, using a two-compartment apparatus; cats showed aversion towards the compartment where full-body restraint occurred (P=0.043; P=0.035). Shelter cats were used to validate behavioural and physiological response differences between passive (n=22) and full-body (n=25) restraint. Full-body restrained cats showed a higher respiratory rate (P=0.004), more lip licks (F1,42 = 6.18; P = 0.017), more side/back ear positions (P<0.0001), and a greater pupil dilation (unfriendly full-body vs unfriendly passive, P=0.0007), than passively restrained cats. These responses were then used to assess cat responses to scruff (n=17), clip (two clips applied to neck skin; n=16), and full-body (negative; n=19), compared to passive restraint. The number of negative responses were highest in full-body (respiratory rate P=0.01; ear P=0.0007, pupil P=0.004, vocalizations P=0.005) and clip (pupil P=0.01, vocalizations P=0.007, ear P=0.02) restrained cats. The results for scruffed cats showed mixed results in comparison to the other restraint groups. This thesis provides the first validated methodologies to assess cat responses to restraint, and suggests that clip restraint may be more negative to scruff restraint, showing a need for further research on alternatives.