Quantitation of the cancer marker LASA using immobilized enzymes in a semiautomated system.

Cancer patients are found to have elevated serum levels of lipid-associated sialic acid (LASA). LASA measurement provides a valuable test for the diagnosis of human cancer. The currently used measurement methods are tedious and nonselective. Here we report a method based on immobilized enzyme minicolumns in a flow system for the analysis of this clinically important analyte. Three enzymes comprising two minicolumns are introduced online and the analyte in a precisely injected volume of 20 microL is made to pass through these immobilized enzyme columns. The final product of the reaction is detected amperometrically. Neuraminidase and NANA-aldolase are coimmobilized on controlled pore glass, followed by a column of pyruvate oxidase. The method was applied to the quantitation of LASA in the sera of normal individuals and patients with cancer of the esophagus. We observed no loss of activity of the immobilized enzymes after analyzing about 300 samples. The sample throughput was 30 samples per hour.