Background There are persistent weaknesses in obtaining accurate reliable and complete data on local and external investments in immunization and yet these are critical to estimate costs, resource needs and gaps so as to aid country financing, planning and resource mobilization. This study aimed to measure and describe the financial envelope for immunization activities and conduct an expenditure analysis at district level. Methods The Systems of Health Accounts (SHA) 2011 methodology was used to quantify and map out the resource envelope for immunization. Data were collected at national and district levels from public and external sources of immunization through key informant interviews coupled with document reviews. Data were coded (SHA), categorized and disaggregated to allow for greater detail on the types of immunization activities. Financing methodology used was largely drawn from Guthrie et al, 2015. Results Over a five-year period, funding for immunization has increased fourfold from US$20.4 million in 2012 to US$ 85.6 million in 2016. The increase in the resource envelope is attributed to the lift of the ban of Gavi funding to Uganda as well as new vaccine introduction. Gavi, the Vaccine Alliance became the biggest contributor (59%) in 2015 and 2016 with 66% of the funds managed by National Medical Stores. Eighty percent of the resources are spent on facility-based routine immunization which includes expenditure on human resources and immunization outreaches. At district level, the overall proportion of total funding to immunization was 15% of their Primary Health Care (PHC) Grant. The bulk (82%) of the PHC funds were spent on supervision to lower health centers (transport and distribution of vaccines) while majority (51%) development partner funds were spent on routine immunization activities (vaccine collection and per-diems/allowances to support outreaches). Conclusion The increasing dependence on development partner support raises issues around sustainability alongside other challenges like misalignment and displacement This warrants an increase in financial commitment to immunization by Government of Uganda, prioritization of resources for immunization at district level, and also operationalization of the existing financial sustainability plan for immunization.