Twenty minutes after the last animal was head-locked, the first basal blood
sample was taken from the coccygeal vein by a single operator. The cows were then
randomly divided into two groups. A second blood sample was taken from the coccygeal vein of animals in the control group (CG, n = 5) after a 20 min rest period. The
cows remained in the head-locks and were approached as calmly as possible by the
same operator in order to minimise stress. The cows to be ‘handled’ (treatment
group [TG], n = 8) were released into an adjacent pen after the first blood sample
was taken, and were subjected to handling procedures by two operators for approximately 8 min. This handling included haltering, driving them back and forth and
walking them back into the first pen, where they were head-locked again. Twenty
minutes after the start of these procedures, a second blood sample was taken from
the TG cows. Overall, this process reflected handling procedures at our research
farm where dairy cows are assembled, driven, head-locked and blood-sampled.