Background: The strip patch test is recommended if patch test results are presumed to be false‐negative. It is under discussion whether the cutaneous inflammation induced by tape stripping is a pivotal element in enhancing the allergic contact dermatitis response. Objectives: To investigate epidermal mRNA expression of mediators implicated in the regulation of inflammation induced by tape stripping according to our standardized protocol in human skin. Patients/Methods: Thirty‐one healthy volunteers (median age 26 years, range 18–62 years) participated in the study. We measured epidermal mRNAs coding for cytokines tumour necrosis factor‐α (TNF‐α), interferon‐γ (IFN‐γ), interleukin (IL)‐1α, IL‐1β, IL‐33, IL‐10, IL‐8/CXCL8, chemokine (C–C motif) ligand 2 (CCL2)/MCP‐1 (monocyte chemotactic protein‐1), chemokine (C–C motif) ligand 5 (CCL5)/RANTES (regulated upon activation normal T‐cell expressed, and presumably secreted), IL‐6, granulocyte–macrophage colony‐stimulating factor (GM‐CSF), an adhesion molecule intercellular adhesion molecule type 1 (ICAM‐1), heat shock proteins (Hsp27, Hsp70 and Hsp90), and the inflammasome complex NACHT‐LRR‐PYD‐containing protein‐3 (NALP3), apoptosis‐associated speck‐like protein containing a CARD (ASC)/CARD5 (caspase recruitment domain‐containing protein‐5) and caspase‐1 in tape‐stripped skin of the upper back relative to untreated skin of the upper back, using quantitative real‐time reverse transcription polymerase chain reaction technology. Results: Epidermal mRNA expression levels of TNF‐α, Hsp90, Hsp70, IL‐33, and IL‐8/CXCL8 were significantly upregulated, whereas CCL5/RANTES expression was significantly downregulated 6 hr after standardized tape stripping. Conclusions: Tape stripping the skin according to our standardized protocol shows immunostimulatory effects, with induction of keratinocyte‐derived ‘alarmins’ and cytokines mounting an immune response, which may contribute to the increased sensitivity of the strip patch test versus the patch test.