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Messenger ribonucleic acid vaccines against coronavirus disease 2019 (COVID-19) have been temporally associated with various renal adverse events, including acute tubulointerstitial nephritis (ATIN). Herein, we present the case of a 14-year-old male who developed biopsy-confirmed ATIN following a booster dose of the BNT162b2 (Pfizer-BioNTech) severe acute respiratory syndrome coronavirus 2 vaccine. Ten days after vaccination, the patient presented with fever, fatigue, and nausea and was referred to our hospital because of impaired kidney function. Kidney biopsy demonstrated diffuse tubulointerstitial inflammation consistent with ATIN, and gallium-67 scintigraphy showed diffuse bilateral kidney uptake. The lymphocyte transformation test (LTT) showed a stronger positive response to the BNT162b2 vaccine than to concomitant medications, suggesting a possible association with vaccination. Corticosteroid therapy led to rapid improvement in kidney function and normalization of tubular injury markers. This case demonstrated that ATIN may rarely occur after booster vaccination, and highlights the potential role of the LTT as a useful tool when multiple possible triggers are present.