Constructing a highly sensitive duplex immunoassay using AuNPs and AgNPs as nanolabels for investigating the epithelial-mesenchymal transition occurring on circulating tumor cells with lung cancer patients

Transformation of epithelial to mesenchymal (EMT) is an important event in the process of tumor initiation, invasion and metastasis. Circulating tumor cells (CTCs) are one kind of important markers in the field of liquid tumor biopsy, whose number and phenotype represent the occurrence and progression of tumors. Therefore, it is our interest to investigate the epithelial mesenchymal transition process occurring on the surface of CTCs. Herein in this work, two proteins of E-cadherin (E-cad) and N-cadherin (N-cad) were selected as representative proteins of EMT process. To achieve simultaneous analysis of E-cad and N-cad on the surface of rare CTCs, we designed a duplex and portable immunosensor using AuNPs and AgNPs as nanolabels to amplify the immunoreaction signals. The dual channel immunosensor not only exhibited good electrochemical responses for recombinant E-cad and N-cad as low as 0.1 ng/mL and 0.05 ng/mL, respectively, but also showed good linear correlations with different numbers of phenotypic CTCs (10-500 cells/10 μL). The above strategy was further employed to inspect the occurrence of EMT on CTCs surface, which displayed a high consistence with other molecular biological characterizations. Finally, this immunoassay was successfully applied to inspect the correlations of numbers, phenotype of CTCs, as well as E-cad and N-cad expressions on these CTCs in bloods of NSCLC patients with disease stage.