Background: Nasal allergen challenge (NAC) is the gold standard for confirming allergen reactivity in allergic rhinitis (AR), especially in complex cases. However, its application is limited due to a lack of standard procedure. The European Academy of Allergy and Clinical Immunology (EAACI) issued guidelines in 2018 to standardize NAC procedure. This study aims to validate the EAACI diagnostic criteria. Methods: 120 adult participants were recruited: 40 with house dust mite-induced AR, 40 with non-allergic rhinitis (NAR), and 40 healthy controls (HC). NAC was performed, and symptoms were evaluated using subjective scales (Total Nasal Symptom Score [TNSS], Linder, Lebel, and Visual Analog score [VAS]) and objective measures (Active Anterior rhinomanometry [AAR], Acoustic Rhinometry [AcRh], and Four-Phase Rhinomanometry[4PR]). A positive NAC diagnosis was defined per EAACI criteria: either a single clearly positive objective or subjective result, or moderate positive findings in one objective and one subjective criterion. Results: NAC sensitivity ranged from 77.5% to 100%; specificity was slightly lower (61.3% to 93.8%). False positives in the HC group were mainly due to clearly positive objective assessments. In the NAR group, false positives resulted from either clearly positive objective changes or concurrent moderate changes in both objective and subjective assessments. Subjective parameters showed higher diagnostic accuracy (area under the curve [AUC] 0.9933–1) compared to objective parameters (AUC 0.7229–0.864). Conclusion: The sensitivity of the EAACI criteria was generally high, albeit with somewhat lower specificity. Relying solely on clearly positive objective parameters or moderate concurrent changes in both objective and subjective measures may lead to false-positive NAC diagnoses. When subjective measures remain negative at sub-maximal allergen concentrations, escalating the concentration is recommended. Among objective assessments, AAR and AcRh are preferred methods.