The aim of this experiment was to determine the effects that positive or negative physical handling has on 5-14 month old non-lactating heifers. The behavioural and physiological responses to the handling were recorded from 48 Holstein-Friesian heifers. The cows were handled twice daily for a 2-5 minute period. During handling the heifers were moved down a 64m route in an outdoor area. Positively handled heifers were patted, stroked, and slowly moved down the corridor. Negatively handled heifers were slapped, hit with a plastic pipe on the rump and were moved quickly down the corridor. The negative handling of the heifers resulted in long duration to approach within 1 or 2m of a person. The negatively handled heifers also had larger flight zones when approached by a human. Whether the person approaching the heifer was familiar or unfamiliar did not have an impact on the amount of time it took the approach a person, however, there was a trend for negatively handled heifers to exhibited an even larger flight zone towards unfamiliar humans. In terms of the physiological response to negative handling, those heifers had higher free total cortisol in the 5 to 15 minute period following the handling event, and also had higher free cortisol levels in the afternoon of a subsequent handling event. This study shows that the manner in how heifers are handled does negatively impact their behaviour (response to humans) and physiology (caused an acute stress response that may become a chronic problem). What is yet to be determined, is the effect that fear brought about by negative handling has on the milk production and welfare of heifers.