Based on the advantages of 1,1’-oxalyldiimidazole chemiluminescence enzyme immunoassay (ODI-CLEIA), two tumor markers, cancer antigen 19-9 (CA 19-9) and carcinoembryonic antigen (CEA), in human serum were simultaneously analyzed using two photomultiplier (PM) tubes connected with a different emission filter such as 523 nm and 615 nm. The sandwich ODI-CLEIA operated using a detection antibody conjugated with horseradish peroxidase (HRP) was able to quantify trace levels of CA 19-9, while the sandwich ODI-CLEIA generated using a detection antibody conjugated with alkaline phosphatase (ALP) analyzed CEA. Capture antibodies of CA 19-9 and CEA were immobilized on the surface of a polystyrene strip well using the interaction between biotin and streptavidin. After washing the well, human serum, containing CA 19-9 and CEA, and two detection antibodies of CA 19-9 and CEA added in the strip well were incubated for 1 hr at room temperature. Then, the mixture of Amplex Red, a substrate of HRP, and fluorescein diphosphate (FDP), a substrate of ALP added in the strip-well, was incubated for 20 min to produce resorufin and fluorescein. After the reaction, ODI-CL reagents (H<sub>2</sub>O<sub>2</sub> and ODI) were added in the well to immediately and simultaneously measure CL emitted from resorufin (615 nm) and fluorescein (523 nm) using the dual PM-tubes. Relative CL intensity was proportionally enhanced with the increase of the tumor marker. We confirmed that the sandwich ODI-CLEIAs operated with the dual PM-tubes can quantify trace levels of CA 19-9 and CEA in a sample with statistically acceptable accuracy, precision, and reproducibility.

Double chemiluminescence enzyme immunoassays capable of simultaneously quantifying CA 19-9 and CEA in a sample

256th National Meeting (2018)

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