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Data on adherence to and acceptability of once daily lamivudine and abacavir are few.Twenty-four U.K. human immunodeficiency virus type-1 infected children 2–13 years of age participated in the Pediatric European Network for the Treatment of AIDS (PENTA) 13 single arm, open label pharmacokinetic study of twice (every 12 hours) versus once (every 24 hours) daily lamivudine and abacavir. Caregivers were asked to complete an adherence questionnaire at screening, week 0 (switch once daily to twice daily) and weeks 4, 12 and 24. Acceptability was also assessed at screening and week 24.Fifteen children were taking lamivudine and abacavir as part of their regimens, 8 lamivudine only and 1 abacavir only. After switching to lamivudine/abacavir every 24 hours, 7 (29%) received once daily regimens for all drugs. Twenty-three (96%) caregivers thought that switching to once daily lamivudine/abacavir would make things a lot/a little easier for their child: 17 (71%) thought it was actually easier after switching. Six mothers with children taking a mixture of twice/once daily drugs changed their mind, whereas all mothers of children on once daily regimens agreed that it was a lot easier. Nonadherence (missing doses in the last 3 days) was reported for 8 of 118 (7%) completed questionnaires; missed doses were reported for every drug in the regimen with reasons such as “not at home,” “forgot” or “routine different from normal.” However, viral loads in all these children remained <100 copies/mL.Adherence to once daily abacavir/lamivudine was good with no evidence of an association between nonadherence and virologic rebound. Acceptability of once daily drugs was best when the whole regimen was dosed once daily.