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Background: Tuberculosis (TB) is the most common opportunistic infection and the leading cause of death in people living with HIV (PLHIV). HIV-infected children are at a higher risk of TB infection and disease compared to those without HIV. Isoniazid preventive therapy (IPT) is an effective intervention in preventing progression of latent TB infection to active TB. The World Health Organization (WHO) currently recommends that all children aged > 12 months and adults living with HIV in whom active TB has been excluded should receive a 6-months course of IPT as part of a comprehensive package of HIV care. Despite this recommendation, the uptake of IPT among PLHIV has been suboptimal globally. This study sought to determine the factors affecting IPT uptake and completion among HIV-infected children in a large HIV care centre in Nairobi, Kenya., Method: This was a cross-sectional mixed methods study comprising of quantitative and qualitative study designs. Medical records of 225 HIV-infected children aged 1 to < 10 years, in care in the Kenyatta National Hospital Comprehensive Care Centre (KNH CCC) were retrospectively reviewed, and 8 purposively selected healthcare providers and 18 consecutively selected caregivers of children were interviewed., Results: IPT uptake among CLHIV in care in the KNH CCC was 68% (152/225) while the treatment completion rate was 82% (94/115). IPT-related health education and counselling were the main facilitators of IPT uptake and completion, while fear of adverse drug reaction, pill burden and lack of an integrated monitoring and evaluation system for IPT were the major barriers., Conclusion: The IPT uptake in this study was low and fell short of the set global target of > 90%. The completion rate was however acceptable. There is an urgent need to address the identified barriers.