Background: Pharmacovigilance of biological medicines is crucial because it ensures that medicines meet the World Health Organization (WHO) standards. In Zambia, there is little information on healthcare professionals' familiarity, knowledge and practices on the pharmacovigilance of biological and biosimilar medicines. Therefore, this study investigated the familiarity, knowledge, and practices related to the pharmacovigilance (PV) of biological and biosimilar medicines at selected hospitals in Lusaka, Zambia. Methods:The study was an analytical questionnaire-based cross-sectional study conducted among healthcare professionals (HCPs) at the Adult hospital, Cancer Diseases hospital, Paediatrics hospital and Women and New Born Hospital in Lusaka. Data were collected over four weeks in May and June 2021 and subsequently analysed using IBM SPSS version 21. The statistical significance was set at a 95% confidence interval. Results: Of 245 participants, only 115 (48.9%) of the HCPs were familiar with biological medicines to a basic understanding. Regarding the term biosimilars, most of the HCPs (40.9%) never heard of this word. The mean score for knowledge regarding the PV consid-How to cite this paper: