The ChAdOx1 nCoV-19 vaccine has been in large-scale use during the COVID-19 pandemic. Limited efficacy compared to mRNA vaccines and certain potential side effects raise the question of whether anti-adenoviral vector antibodies influence immune responses against the vaccine. Complement activation by ChAdOx1 and leukocyte phagocytosis of ChAdOx1 in vitro were studied. Plasma IgG levels against ChAdOx1 and human adenovirus 2 (hAdV2) hexon protein were determined (n = 20) and IgGs from high- and low-titre plasmas were isolated (n = 3). Complement activation was measured as cleavage of C3 by immunoblotting and generation of C3a and sC5b-9 by ELISA. pHrodo-labelled ChAdOx1 was opsonized with complement and IgG, and phagocytosis by isolated blood PMNs in vitro was studied by flow cytometry. The transcriptomic profile of PMN cells exposed to ChAdOx1 was analysed by RNA-seq. ChAdOx1 activated the classical complement pathway in an anti-adenovirus antibody-dependent manner. Generation of the terminal complement complex sC5b-9 in individual sera correlated with anti-hAdV2 hexon and anti-ChAdOx1 IgG levels. Phagocytosis of ChAdOx1 also correlated significantly with anti-hAdV2 hexon IgG, anti-ChAdOx1 IgG and serum sC5b-9 levels. High-titre anti-hAdv2 hexon IgG increased phagocytosis in the presence of normal serum. Anti-vector antibodies induced rapid complement activation and promoted phagocytosis of the ChAdOx1 vaccine by neutrophils. Moreover, transcriptomic analysis revealed upregulation of complement-related genes induced by the ChAdOx1 vaccine in vitro. Anti-adenovirus vector antibodies and complement activation may thus influence the efficacy of the ChAdOx1 vaccine against SARS-CoV-2 and be also involved in vaccine-related side effects.