Comparison of woolly mammoth (left) and American mastodon

At the beginning of the Pleistocene, elephantids experienced a high rate of speciation. Loxodonta atlantica became the most common species in northern and southern Africa but was replaced by Elephas iolensis later in the Pleistocene. Only when Elephas disappeared from Africa did Loxodonta become dominant once again, this time in the form of the modern species. Elephas diversified into new species in Asia, such as E. hysudricus and E. platycephus; the latter the likely ancestor of the modern Asian elephant. Mammuthus evolved into several species, including the well-known woolly mammoth. In the Late Pleistocene, most proboscidean species vanished during the Quaternary glaciation which killed off 50% of genera weighing over 5 kg (11 lb) worldwide.[39] The Pleistocene also saw the arrival of Palaeoloxodon namadicus, the largest terrestrial mammal of all time.

Proboscideans experienced several evolutionary trends, such as an increase in size, which led to many giant species that stood up to 5 m (16 ft) tall. As with other megaherbivores, including the extinct sauropod dinosaurs, the large size of elephants likely developed to allow them to survive on vegetation with low nutritional value. Their limbs grew longer and the feet shorter and broader. Early proboscideans developed longer mandibles and smaller craniums while more advanced ones developed shorter mandibles, which shifted the head's centre of gravity. The skull grew larger, especially the cranium while the neck shortened to provide better support for the skull. The increase in size led to the development and elongation of the mobile trunk to provide reach. The number of premolars, incisors and canines decreased. The cheek teeth (molars and premolars) became larger and more specialized, especially after elephants started to switch from C3-plants to C4-grasses, which caused their teeth to undergo a three-fold increase in teeth height as well as substantial multiplication of lamellae after about five million years ago. Only in the last million year or so did they return to a diet mainly consisting of C3 trees and shrubs. The upper second incisors grew into tusks, which varied in shape from straight, to curved (either upward or downward), to spiralled, depending on the species. Some proboscideans developed tusks from their lower incisors. Elephants retain certain features from their aquatic ancestry, such as their middle ear anatomy and the internal testes of the males.

There has been some debate over the relationship of Mammuthus to Loxodonta or Elephas. Some DNA studies suggest Mammuthus is more closely related to the former while others point to the latter. However, analysis of the complete mitochondrial genome profile of the woolly mammoth (sequenced in 2005) supports Mammuthus being more closely related to Elephas. Morphological evidence supports Mammuthus and Elephas as sister taxa while comparisons of protein albumin and collagen have concluded that all three genera are equally related to each other. Some scientists believe a cloned mammoth embryo could one day be implanted in an Asian elephant's womb.