Paired acute and baseline serum or plasma tryptase sampling and determination have recently been included as a mechanistic approach in the diagnostic and management guidelines of perioperative immediate hypersensitivity and anaphylaxis. The timing of this paired sampling is clearly defined in international consensus statements, with the optimal window for acute tryptase sampling between 30 minutes and 2 hours after the initiation of symptoms, whereas baseline tryptase should be measured in a sample collected before the event (preop) or at least 24 hours after all signs and symptoms have resolved. A transient elevation of the acute tryptase level greater than [2 + (1.2 × baseline tryptase level)] supports the involvement and activation of mast cells. Here, we provide the clinical, pathophysiological, and technical rationale for the procedure and interpretation of paired acute and baseline tryptase. Clinical examples, up-to-date knowledge of hereditary α-tryptasemia as a frequent cause of baseline tryptase of 7 μg/L and higher, mastocytosis, other clonal myeloid disorders, cardiovascular or renal failure, and technical improvements resulting in continued lowering of the 95th percentile value are discussed. Clues for improved management of perioperative immediate hypersensitivity and anaphylaxis include (1) sustained dissemination and implementation of updated guidelines; (2) preoperative sample storage for deferred analysis; (3) referral for thorough allergy investigation, screening for mast cell–related disorders, and recommendations for future anesthetic procedures; and (4) sustained collaboration between anesthesiologists, immunologists, and allergists.