Our objective was to investigate the effect of overstocking the feed bunk on dairy cow behavioral responses to human approach and reactivity to blood sampling. One hundred and 20 dry Holstein cows were allocated to 1 of 2 treatment groups with different stocking densities [overstocked (OS): 0.88 headlocks/cow; understocked (US): 1.17 headlocks/cow]. Over 2 testing periods (7 d apart), flight response was assessed using a human-approach test with a 5-point ordinal scale defining the distance at which the cow stepped away from the approaching experimenter (0 = not approachable from 3 m to 4 = cow moves away when experimenter is 0 m from the cow). A qualitative assessment was also made of the cow’s response to the experimenter using a visual analog scale (VAS) that included the terms: relaxed, nervous, alert, shy, aggressive, social, and curious. Reactivity to blood sampling via the coccygeal vein was assessed in the pen using a 4-point scale (0 = least reactive to 3 = most reactive). Data were analyzed through a mixed model analysis, using treatment, time, and their interaction. The relationship between qualitative measures was assessed using a Pearson correlation. Treatment did not affect the cow’s flight response; however, there was a significant treatment by time interaction whereby flight response scores decreased with time in OS cows and increased with time in US cows (OS: 1.65 to 1.47, US: 1.33 to 1.68; P = 0.02). Reactivity to blood sampling did not differ by treatment (OS: 1.11, US: 0.98; P = 0.47), and there was also no treatment by time interaction (OS: 1.17 to 1.11, US: 1.01 to 0.98; P = 0.88). The overall correlation between qualitative terms was low. However, the terms ‘relaxed’ and ‘nervous’ showed a significant negative correlation (OS: r = −0.71; P < 0.0001; US: r = −0.61; P < 0.0001). In conclusion, overstocking the feed bunk affected the animal’s response to an approaching human. Cows in the OS treatment became less approachable over time, which may indicate fear, stress, or an increase in arousal. Future research should investigate the effect overstocking may have on cow temperament for a longer duration, as this may further decrease approachability.