It has been suggested that true vipers (Viperidae: Viperinae) possess the ability to detect temperature differences between objects despite the lack of an apparent infrared radiation sensor. We tested the ability to distinguish between heated and unheated targets in three species of pitvipers (Viperidae: Crotalinae), four species of true vipers, two species of colubrids (Colubridae: Natricinae, Colubrinae) and Azemiops feae (Viperidae: Azemiopinae). All species of pitvipers tested could distinguish between the warm and cool targets, while no tested species of true viper, colubrid or Azemiops demonstrated this ability. In addition, pitvipers exhibited behaviors that true vipers or Azemiops did not exhibit. Our results suggest that the tested species of true vipers, Azemiops and colubrids may not posses the ability to sense infrared radiation or do not use it in a defensive context, and suggest that some defensive behaviors are associated with the pit organ in pitvipers.