The study investigated equine responses to novelty and handling, aiming to reveal whether horse–human relationships reflect criteria of an attachment bond. Twelve adult Standardbreds were subjected to a fear-eliciting test (novel objects presented close to two humans) and a handling test (being led passing novel objects) to study attachment-related behaviours and ease of handling. The tests were performed both before (pre-test) and after (post-test) horses had been trained by the same female handler (10 sessions of 15 min). Horses were assigned to three groups of four, each of which underwent different operant conditioning protocols: negative reinforcement (NR; pressure, release of lead, and whip tap signals) or combined NR with either positive reinforcement using food (PRf) or wither scratching (PRs). Results showed that neither familiarity of the person nor training method had a significant impact on the horses’ behavioural responses in the post-tests. However, horses showed decreased heart rates between pre-and post-tests, which may indicate habituation, an effect of training per se, or that the presence of the familiar trainer served to calm the horses during the challenging situations. There were large individual variations among the horses’ responses and further studies are needed to increase our understanding of horse–human relationships. © 2021 by the authors.