Development of a quantum-dot-labelled magnetic immunoassay method for circulating colorectal cancer cell detection

Figure 1 Circulating tumour cells are separated from the leucocytes using magnetic beads coupled with epithelial cell adhesion molecule antibody and a magnetic device. These complexes are then tagged with cytokeratin-19 (CK19) biotinylated antibody and streptavidin-conjugated quantum dots (QDs) which lead to the detection of a fluorescent signal. EpCAM: Epithelial cell adhesion molecule; DLD-1: Human colon adenocarcinoma cell line.

Figure 2 Representative fluorescence-activated cell sorting analysis of peripheral blood samples. A: Events which fell within region R1 are confirmed to be cytokeratin 19 (CK19)+ and fluorescent in FL3 (anti-CK19 conjugated with QDs); B: From the R1 gate, the events confirmed to be CK19+CD45-are those in the UL region, in UR region the events represent those that are CK19+CD45+ and fluorescent in FL1 (anti-CD45 conjugated with Alexa Fluor). We deemed that the cells in the UL are those meeting the criteria for circulating tumor cells.

Figure 3 Representative results recorded by the proposed method for the human colon adenocarcinoma cell line serial dilutions ranging from 10⁴ to 10 cells/mL (numbers 1-5) and negative controls. Fluorescence is evident in the human colon adenocarcinoma cell line samples with concentration up to 10 cells/mL, as illustrated at the fluorescent emission spectra obtained after magnetic separation.