Antibody use is a critical component of cardiovascular physiology research, and antibodies are used to monitor protein abundance (immunoblotting) and protein expression and localization (in tissue by immunohistochemistry and in cells by immunocytochemistry). With ongoing discussions on how to improve reproducibility and rigor, the goal of this review is to provide best practice guidelines regarding how to optimize antibody use for increased rigor and reproducibility in both immunoblotting and immunohistochemistry approaches.