The assay could be used for simultaneously detecting three lung cancer biomarkers: carcinoembryonic antigen (CEA), fragments of cytokeratin 19 (CYFRA21-1) and NSE in 10 μl of human serum, with a wide linear dynamic range (1.03–111 ng/ml for CEA and CYFRA21-1; 9.26–1000 ng/ml for NSE) and a low detection limit (CEA: 0.19 ng/ml; CYFRA21-1: 0.97 ng/ml; NSE: 0.37 ng/ml; S/N = 3). This evidence concerns the gene ENO2 and lung cancer.