The popliteal lymph node (PLN) assay has been proposed as a tool to predict in rodents those xenobiotics likely to induce autoimmune reactions in humans. To further validate this assay and to study the mechanisms involved, histologic changes in PLNs from rats injected with streptozotocin, diphenylhydantoin, pure acetone, or 50% ethanol were compared to a local graft‐versus‐host (GvH) reaction. This study suggests that routine histology of PLNs is instrumental to discard primary irritants. In addition, the hypothesis of a CvH‐like mechanism in positive PLN responses is supported by the finding that the reference compounds streptozotocin and diphenylhydantoin produced histologic changes similar to a “true” local GvH reaction.