BACKGROUND Allergy to penicillins remains an important issue. Penicillin skin testing (PST) with major and minor determinants has been shown to be a highly valuable tool for identifying IgE-mediated penicillin allergy. The value of additional testing with side-chain-specific moieties from semisynthetic penicillins such as amoxicillin is not well-established in spite of the widespread use of these medications. METHODS A retrospective review of all consecutive inpatient PST results from 1995 to 2007 comprising 1,068 patients was performed in our institution on individuals with a self-reported history of beta-lactam allergy to assess the importance of including the amoxicillin determinant in a previously validated PST panel. Descriptive statistics were performed. The PST panel included penicilloyl-polylysine, penicillin G, penicilloate, penilloate and amoxicillin. RESULTS Of 1,068 patients, 243 (23%) had a positive skin test reaction on the PST panel. Testing with amoxicillin was positive in 30.9% of patients, the majority of whom (81%) were also positive to 1 or more standard penicillin reagents. Fourteen of the 243 positive patients (5.8%) had a positive skin test reaction only to amoxicillin. Additionally, the use of penicilloate and penilloate minor determinants in combination with penicillin G identified a greater percentage of penicillin-allergic individuals compared to using only penicillin G (22.6 vs. 6.6%), demonstrating their importance in the PST panel. CONCLUSIONS These data indicate that the inclusion of the amoxicillin determinant appears to identify a small but important group of allergic individuals who may otherwise test negative on a PST panel.