The generation of monoclonal antibodies against human pancreatic exocrine cancer: A study of six different immunisation regimes

Abstract

Six different immunisation regimes have been used to generate spleen cells with reactivity against human pancreatic exocrine cancer. Immunised spleen cells were fused with an NSO/1 myeloma line and supernatants from these hybridomas selectively screened for monoclonal antibodies which bound predominantly to a pancreatic cancer cell line (GER). The spleen cells from hairy litter mates immunised with pancreatic cancer xenograft homogenates and viable GER cells generated 13% of hybridoma supernatants which showed some selectivity for GER pancreatic cancer cells in a fixed cell ELISA assay. The other methods produced only 4% of hybrids with selectivity for GER cells. The antigen distribution on gluteraldehyde fixed cells was similar to that found for viable cell monolayers but many antigens were unstable on formalin fixation. Immunohistochemical staining of GER cells grown on glass slides showed a heterogeneity of antigen distribution with up to 70% of the cells exhibiting a vesicular pattern of staining. Fifty percent of the antibodies which bound to GER cells were also reactive against antigens in formalin-fixed paraffin-embedded tissue sections of the original GER tumour. Monoclonal antibody DD9E7 identified an antigen expressed on 12/14 pancreatic adenocarcinomas. The antibody showed strong staining of malignant luminal membranes and cytoplasm. The antigen was also present in normal salivary and sweat glands, and colon and breast carcinomas, but its tissue distribution was unlike that of CEA or EMA. The expression of this antigen in 12/14 of pancreatic carcinomas suggests that DD9E7 may be a useful reagent for pancreatic tumour detection. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6

The immunoperoxidase localization of tumour markers in ovarian cancer: the value of CEA, EMA, cytokeratin and DD9

“…Where patients had bilateral ovarian neoplasms they were of the same histological type, and in all six patients both tumours either expressed or did not express CEA and DD9. Some of the tumours, particularly serous carcinomas, contained numerous granulocytes and/or macrophages which made interpretation of DD9 staining, present in these normal cells (Grant et al 1985), more difficult (Fig. 8).…”

“…Sequential 4 μm sections of one representative block of each of the tumours were immunostained using an indirect immunopcroxidase technique (Heyderman 1983). The primary antibodies used were mouse monoclonal antibodies to CEA (Amersham International plc, Bucks), EMA (Dako Ltd, Bucks), CAM 5.2 (Beckton Dickinson, Oxford) and DD9‐E7 (Grant et al 1985). The secondary antibody was an affinity purified sheep anti‐mouse immunopcroxidase conjugate (Amersham).…”

“…We decided to investigate the immunocytochemical expression of carcinoembryonic antigen (CEA) (Gold & Freedman 1965), epithelial membrane antigen (EMA) (Heyderman et al 1979), cytokeratin (CAM 5.2) (Makin et al 1984), and DD9 (Grant et al 1985) to determine whether the expression of these markers might be of prognostic or diagnostic value.…”

“…The monoclonal antibody DD9‐E7 was raised against the GER pancreatic exocrine carcinoma cell line (Grant et al 1985). The epitope to which the antibody binds is expressed by all pancreatic adenocarcinomas studied by us (Heyderman et al 1990) and by a variety of other epithelial neoplasms.…”

Purification and characterization of a peanut‐agglutinin‐binding pancreatic‐cancer‐related serum mucus glycoprotein

“…Several tumour markers have been reported in association with exocrine pancreatic cancer (Banwo et al, 1974;Chu et al, 1977;Gelder et al, 1978;Mihas, 1978;Schultz and Yu-N s , 1979;Kuntz and Archer, 1979;Koprowski et al, 1979;Shimano et al, 1981;Metzgar et al, 1984;Chin and Miller, 1985;Grant et al, 1985;Yuan et al, 1985;Ohyanagi et al, 1987;Kajiji et al, 1987), a cancer that is increasingly common and notoriously difficult to diagnose at an early stage. These tumour markers have generally been detected by the development of polyclonal or monoclonal antibodies against extracts of tumours or tumour-cell lines, some of which were not pancreatic in origin.…”