The objective of this study was to evaluate, over 2 consecutive years, the effects of acclimation to human interaction on performance, temperament, plasma concentrations of hormones and metabolites, and pregnancy rates of Brahman-crossbred cows. A total of 160 Braford and 235 Brahman x British cows were assigned to the 2-yr study. Approximately 45 d after weaning (August 2006) in yr 1, cows were evaluated for BW, BCS, and temperament (chute score, pen score, and exit velocity), stratified by these measurements in addition to breed and age, and randomly allocated to 14 groups (Braford = 8; Brahman x British = 6). Groups were randomly assigned to the control or acclimation treatment. In yr 2, cows were reevaluated within 45 d after weaning (August 2007) for BW, BCS, and temperament, stratified, and divided into 14 groups similarly as in yr 1, but in such a way that cows received the same treatment assigned in yr 1. Cows were acclimated to human interaction from August to January, and the acclimation process consisted of the same person visiting groups twice weekly and offering approximately 0.05 kg of range cubes per cow (as-fed basis). In January of both years, cow temperament, BW, and BCS were reassessed and cows were exposed to a 90-d breeding season. Blood samples were collected at the beginning of the acclimation period (August) and breeding season (January) for determination of plasma cortisol, IGF-I, and acute phase proteins. A treatment x breed interaction was detected during yr 1 (P < 0.01) for pregnancy analysis because acclimated Braford cows conceived earlier and at a greater percentage (P < 0.01) compared with control cows. According to values obtained at the beginning of breeding and pooled across treatments and breeds, IGF-I concentrations and BCS affected quadratically (P < 0.05), and concentrations of ceruloplasmin and haptoglobin decreased linearly (P < 0.05), the probability of pregnancy during both years. Temperament and cortisol concentrations decreased the probability of pregnancy linearly (P < 0.05) during yr 1 (only Braford cows for cortisol analysis) and affected the probability of pregnancy quadratically (P < 0.05) during yr 2. Results from this study indicate that acclimation did not affect cow temperament and physiological responses but did increase pregnancy rates of Braford cows during yr 1. Further, measurements and physiological responses associated with temperament influenced the probability of cows becoming pregnant during the breeding season.