A NEW METHOD FOR DETECTING BETA-1,4-GALACTOSYLTRANSFERASE ACTIVITY INSERA OF CANCER-PATIENTS

A new method for assaying the activity of the enzyme that catalyzes th e formation of a cancer-associated glycolipid, paragloboside (nLc(4)Ce r), from lactotriaosyl-ceramide (Lc(3)Cer) and UDP-galactose has been developed that is based on a time-resolved fluoroimmunoassay (TRFIA) w ith a Europium (Eu)-chelate-labeled antibody. The substrate, Lc(3)Cer, immobilized on a microtiter plate, was incubated with UDP-galactose, MnCl2, Triton CF-54, and the enzyme. The content of the incubation pro duct, nLc(4)Cer, was determined by the TRFIA with anti-nlc(4)Cer monoc lonal antibody H-11 as the first antibody and Eu-labeled anti-mouse Ig M antibody as the second one. The lower limit of detection of nLc(4)Ce r was estimated to be 0.2 pmol. This method was used to detect the gal actosyltransferase activity in sera from patients with colorectal canc er or benign colorectal adenomas and from healthy subjects of a refere nce sample group. The reference interval was 0-0.25 pmo1/25 mu l serum /2 h. Activity was significantly greater in patients with colorectal c ancer than in those with colorectal benign adenoma (P < 0.05) and the subjects of the reference sample group (P < 0.01). (C) 1994 Academic P ress, Inc.