BACKGROUND: Rapid detection of influenza infection is important for patient management and timely anti-viral therapy. Rapid antigen detection tests for influenza have inferior sensitivity when compared to nucleic acid-based amplification tests. An isothermal nucleic acid amplification test that offers the potential for rapid molecular testing at the clinical point-of-care with simple equipment can improve influenza detection rates. OBJECTIVES: To evaluate the performance of Alere i Influenza A&B isothermal nucleic acid amplification test to detect influenza A and B in comparison to viral cell culture as reference method. STUDY DESIGN: A prospective, multicenter, clinical study to evaluate the clinical performance of the Alere i Influenza A&B assay in a point-of-care setting using prospectively enrolled specimens from both children and adults was conducted in seven sites. RESULTS: In comparison with viral cell culture, the overall sensitivity and specificity of the Alere i Influenza A&B assay was 97.8% and 85.6% for the detection of influenza A, and 91.8% and 96.3% for the detection of influenza B, respectively. Following resolution of discrepant results by real-time RT-PCR the sensitivity and specificity of the Alere i Influenza A&B assay improved to 99.3% and 98.1% for influenza A, and 97.6% and 100% for influenza B, respectively. CONCLUSIONS: The Alere i Influenza A&B isothermal nucleic acid amplification test is an ideal point-of-care test for influenza detection in children and adults due to its high sensitivity and specificity and ability to generate results within 15min from specimen receipt.