Leaves from cotton, Gossypium hirsutum L., plants containing transgenic traits (Bollgard®, Bollgard II®, and WideStrike™) were assayed for bioactivity against bollworm, Helicoverpa zea (Boddie); beet armyworm, Spodoptera exigua (Hübner); fall armyworm, Spodoptera frugiperda (J. E. Smith); and cabbage looper, Trichoplusia ni (Hübner); in a laboratory. Fifty leaves from the middle of the plants were collected every 20 days, starting at 40 days and until 120 days after cottons were planted. The leaves were fed to 15 first-instar larvae of each insect pest until they died or pupated. Most larvae died when fed Bollgard II or WideStrike. At the end of the season (100–120 days after planting), more larvae died when fed Bollgard II than WideStrike. Conclusions from the results are that (1) mortality was not significantly different when the larvae were fed dual Bt cotton leaves of different ages; (2) survival duration depended on consumption of the amount of endotoxin that caused larval mortality; (3) biological characteristics of surviving lepidopterans (pupal weight, emergence, and developmental time) were significantly better on non-Bt cotton.