A new CEA-related substance, (CEA_low) was isolated from liver metastases of colorectal cancer. Purified CEAlow was homogeneous by several physico-chemical criteria. It had an apparent molecular weight of 125,000. 2/9 monkey anti-CEA sera gave a spur with CEA over CEA_low in immunodiffusion. Analysis of peptide relatedness showed that CEA_low was more closely related to CEA than two other purified CEA related antigens, nonspecific cross-reacting antigen (NCA) and biliary glycoprotein I (BGPI). Since CEA_low appeared to lack unique determinants, we tentatively conclude that it is a naturally occuring fragment of CEA.

Of seven monoclonal antibodies (mabs) produced against CEA, all reacted with CEA_low, 5 reacted strongly with NCA-2 (a highly cross-reactive meconium antigen) while 2 mabs were only weakly reactive, 2 were reactive with NCA and 1 gave a weak reaction with BGPI. Five different epitopes present in the peptide moiety of CEA were recoonized by these mabs. Affinity constants of 1.2-7.4x10⁸M^-T were obtained. Two mabs bound to >2 epitopes on CEA indicating regions of homology in CEA.

Three mabs were used for tumor immunolocalization of CEA, containing human tumor cells (LS174T, Detroit 562 and HT29) grafted in nude mice. Excellent tumor localization was obtained with one radiolabelled antibody. A second antibody gave an intermediary degree of tumor localization while a third mab was not enriched in the tumor. Normal mouse IgG did not localize in the tumor. Similarly anti-CEA mab did not bind to grafts of a non-CEA producing sarcoma. By y-scintigraphy, the first antibocty gave a definite spot corresponding to the tumor.

Two mabs were used as reagents in a two site monoclonal enzyme immunoassay (MEIA) for CEA. Both were nonreactive with NCA and BGPI. One mab was strongly reactive with NCA-2 while the other was only weakly reactive with this antigen. Interestingly, in the MEIA, all normal CEA-related antigens, including NCA-2, and normal tissue extracts were nonreactive. Sera from healthy individuals gave values <5 ug CEA/1 with this MEIA and with a RIA tested in parallel. However, the CEA values in patients with nonmalignant liver disease and ulcerative colitis were significantly lower in the MEIA than in the RIA. No difference in CEA values between the two assays was seen in patients with colon-, lung-, pancreas and breast carcinoma. We conclude that this MEIA has an increased specificity for carcinoma than conventional immunoassays against CEA.