Background Tuberculosis (TB) remains a major health problem in Zambia, despite considerable efforts to control and prevent it. With this study, we aim to understand how perceptions and cultural, social, economic, and organisational factors influence TB patients’ pre-hospital delay and non-compliance with care provided by the National Tuberculosis Programme (NTP). Methods A mixed methods study was conducted with 300 TB patients recruited at Kanyama clinic for structured interviews. Thirty were followed-up for multiple in-depth interviews. Six focus group discussions were organised and participant observation was conducted. Ten biomedical care providers, 10 traditional healers, and 10 faith healers were interviewed. Factors associated with non-compliance (disruption of treatment > one week) were assessed by applying logistic regression analyses; qualitative analysis was used to additionally assess factors influencing pre-hospital delay and for triangulation of study findings. Results TB treatment non-compliance was low (10 %), no association of outcome with cultural or socio-economic factors was found. Only patients’ time constraints and long distance to the clinic indicated a possible association with a higher risk of non-compliance (OR 0.52; 95 % CI 0.25, 1.10, p = 0.086). Qualitative data showed that most TB patients combined understandings of biomedical and traditional TB knowledge, used herbal, traditional and/or faith healing, suffered from stigmatizing attitudes, experienced poverty and food shortages, and faced several organisational obstacles while being on treatment. This led in some cases to pre-hospital delay or treatment non-compliance. Conclusions Mixed methods analysis demonstrated the importance of in-depth information ascertained by qualitative approaches to understand how cultural, socio-economic and organisational factors are influencing patients’ pre-hospital delay and treatment compliance. To strengthen the Zambian NTP, combating stigma is of utmost priority coupled with programmes addressing poverty. Organisational barriers and co-operation between (private) clinics and traditional/faith healers should be considered. Electronic supplementary material The online version of this article (doi:10.1186/s12889-016-3771-9) contains supplementary material, which is available to authorized users.