The epidermal growth factor (EGF) receptor and its ligands are crucially involved in the renal response to ischaemia. We studied the heparin binding‐epidermal growth factor (HB‐EGF), a major ligand for the EGF receptor, in experimental and human ischaemia/reperfusion injury (IRI). HB‐EGF mRNA and protein expression was studied in rat kidneys and cultured human tubular (HK‐2) cells that were subjected to IRI and in human donor kidneys during transplantation. The effect of EGF receptor inhibition was investigated in vivo and in vitro. Furthermore, urinary HB‐EGF protein excretion was studied after renal transplantation. Finally, HB‐EGF KO and WT mice were subjected to IRI to study the role of HB‐EGF in renal injury. HB‐EGF mRNA was significantly up‐regulated in the early phase of IRI in rats, cells, and human donor biopsies. Treatment with PKI‐166 reduces macrophage accumulation and interstitial α‐SMA in the early phase of IRI in rats. In vitro, PKI‐166 causes a marked reduction in HB‐EGF‐induced cellular proliferation. Urinary HB‐EGF is increased after transplantation compared with control urines from healthy subjects. HB‐EGF KO mice subjected to IRI revealed significantly less morphological damage after IRI, compared with WT mice. We conclude that IRI results in early induction of HB‐EGF mRNA and protein in vivo and in vitro. Absence of HB‐EGF and inhibition of the EGF receptor in the early phase of IRI has protective effects, suggesting a modulating role for HB‐EGF. Copyright © 2010 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.