Drug‐induced liver injury (DILI) frequently has a delayed onset with several human leukocyte antigen (HLA) genotypes affecting susceptibility, indicating a potential role for the adaptive immune system in the disease. The aim of this study was to investigate whether drug‐responsive T lymphocytes are detectable in patients who developed DILI with the combination, antimicrobial amoxicillin‐clavulanate. Lymphocytes from 6 of 7 patients were found to proliferate and/or secrete interferon‐gamma (IFN‐γ) when cultured with amoxicillin and/or clavulanic acid. Amoxicillin (n = 105) and clavulanic acid (n = 16) responsive CD4⁺ and CD8⁺ T‐cell clones expressing CCR, chemokine (C‐C motif) receptor 4, CCR9, and chemokine (C‐X‐C motif) receptor 3 were generated from patients with and without HLA risk alleles; no cross‐reactivity was observed between the two drug antigens. Amoxicillin clones were found to secrete a heterogeneous panel of mediators, including IFN‐γ, interleukin‐22 and cytolytic molecules. In contrast, cytokine secretion by the clavulanic acid clones was more restricted. CD4⁺ and CD8⁺ clones were major histocompatability complex class II and I restricted, respectively, with the drug antigen being presented to CD4⁺ clones in the context of HLA‐DR molecules. Several pieces of evidence indicate that the clones were activated by a hapten mechanism: First, professional antigen‐presenting cells (APCs) were required for optimal activation; second, pulsing APCs for 4‐16 hours activated the clones; and third, inhibition of processing abrogated the proliferative response and cytokine release. Conclusion: Both amoxicillin‐ and clavulanic acid–specific T cells participate in the liver injury that develops in certain patients exposed to amoxicillin‐clavulanate. (Hepatology 2015;62:887‐899)