<jats:title>Abstract</jats:title><jats:p>Intraindividual tumoural heterogeneity (ITH) is a hallmark of solid tumours and impedes accurate genomic diagnosis and selection of proper therapy. The aim of this study was to identify ITH of ovarian high‐grade serous carcinomas (OSCs) and to determine the utility of ascitic cancer cells as a resource for mutation profiling in spite of ITH. We performed whole‐exome sequencing, copy number profiling and DNA methylation profiling of four OSC genomes by using multiregional biopsies from 13 intraovarian lesions, 12 extraovarian tumour lesions (omentum/peritoneum), and ascitic cells. We observed substantial levels of heterogeneity in mutations and copy number alterations (CNAs) of the OSCs. We categorized the mutations into ‘common’, ‘shared’ and ‘private’ according to the regional distribution. Six common, eight shared and 24 private mutations were observed in known cancer‐related genes. Common mutations had a higher mutant allele frequency, and included <jats:italic>TP53</jats:italic> mutations in all four OSCs. Region‐specific chromosomal amplifications and deletions involving <jats:italic>BRCA1</jats:italic>, <jats:italic>PIK3CA</jats:italic> and <jats:italic>RB1</jats:italic> were also identified. It is of note that the mutations detected in ascitic cancer cells represented 92.3–100% of overall somatic mutations in the given case. Phylogenetic analyses of ascitic genomes predicted a polyseeding origin of somatic mutations in ascitic cells. Our results demonstrate that, despite ITH, somatic mutations, CNAs and DNA methylations in both ‘common’ category and cancer‐related genes were highly conserved in ascitic cells of OSCs, highlighting the clinical relevance of genome analysis of ascitic cells. Ascitic tumour cells may serve as a potential resource for discovering somatic mutations of primary OSC with diagnostic and therapeutic relevance. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.</jats:p>