Most spiders have venom glands. Only spiders of the Uloboridae and Holoarchaeidae families have no venom. The venom glands lie either in the chelicerae or at the front of the cephalothorax. The venom duct leads to the fangs, which are located at the end of the chelicerae. The venom gland consists of an outermost muscle layer, an underlying secretion layer and a duct. The venom is injected into the natural prey but is also used to defend against predators. The venom glands probably developed over the course of evolution from modified salivary glands whose secretions helped in the extracorporeal digestion of the prey. After having killed the prey, the spider releases digestive juices over it and afterwards sucks up the liquefied mush. In spiders of the suborder of the Orthognata (e.g. Atrax, Hadronyche, Aphonopelma, Trechona, Dugesiella) the chelicerae move forwards-downwards, which means that these spiders must lift themselves somewhat to take a downwards bite. The chelicerae move diagonally in the Labidognatha (Latrodectus, Loxosceles, Phoneutria, Cheiracanthium).

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Spider venom has various purposes. Spiders which hunt, such as Loxosceles and Phoneutria, have neurotoxic / proteolytic toxins in their venom. Spiders which make webs generally have weaker venom, except for Latrodectus. Some bird spiders have urticarial hairs which, when lost by the animal, can irritate skin, conjunctivae and the mucous membrane of the mouth. Similar irritating hairs are also found in other animals, such as some caterpillars (e.g. procession caterpillar) and various adult butterflies. The irritating hairs are also present on the cast-off skin (exuvia) when the animal moults.