Monoclonal Antibodies Reactive with the Breast Carcinoma-Associated Mucin Core Protein Repeat Sequence Peptide Also Recognise the Ovarian Carcinoma-Associated Sebaceous Gland Antigen

Layton, Guy T.; Devine, Peter L.; Warren, J; Birrell, Geoff W.; Xing, Puyuan; Ward, Bruce G.; McKenzie, Ian F. C.

doi:10.1159/000217661

Cited by 29 publications

(21 citation statements)

References 14 publications

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“…The LS1 74T and HT29-SB colon cancer cell lines have been shown to secrete significant quantities of mucin (Devine et al, 1991 (Layton et al, 1990). Positive clones were then checked by ELISA on Ml, and by immunoblotting on LS174T and HT29-SB mucins .…”

Section: Mucinsmentioning

confidence: 99%

“…Inhibition ELISA Optimal concentrations of antigens and MAbs were determined by checkerboard ELISA, and inhibition ELISA was performed (Layton et al, 1990). Briefly, MAbs at twice the required concentration (3A2 ascites 2000-', 4F1 ascites 5000-1) were incubated for 3 h at RT with an equal volume of peptide (64 fM) in PBS-Tween.…”

Section: Elisamentioning

confidence: 99%

“…Five distinct gene loci have now been identified in humans, these being renamed MUC1, MUC2 and MUC3 (Human Gene Mapping Nomenclature Committee, 1989) and the names MUC4 and MUC5 have been proposed Aubert et al, 1991 (Gendler et al, 1987;Gum et al, 1989;Porchet et al, 1991;Aubert et al, 1991). Many monoclonal antibodies (MAbs) reacting with the MUC1 VNTR have been reported (Gendler et al, 1988;Xing et al, 1990;Layton et al, 1990;Price et al, 1990), and assays incorporating some of these MAbs have been shown to be particularly useful in monitoring patients with breast and ovarian cancer (Ward et al, 1993;Safi et al, 1991;Bhargava et al, 1989). As well as overexpression of MUC1, altered glycosylation of the VNTR is responsible for the exposure of these peptide epitopes in tumours (Gendler et al, 1988;Devine et al, 1990a).…”

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confidence: 99%

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Monoclonal antibodies reacting with the MUC2 mucin core protein

Pl¹,

McGuckin²,

Birrell³

et al. 1993

Br J Cancer

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Summary This study sought to produce monoclonal antibodies (MAbs) which reacted with the MUC2 core protein. Two MAbs [3A2 (IgGl) and 4F1 (IgM)] were produced by immunising female BALB/c mice with gel-formed mucin from the LS174T colon cancer cell line followed by a KLH conjugate of a 29 amino acid synthetic peptide whose sequence was derived from the variable number of tandem repeats (VNTR) region of a MUC2 cDNA clone.The MAbs reacted with synthetic MUC2 VNTR peptides but not synthetic MUCI or MUC3 VNTR peptides, and showed specific reactivity in Western blotting with a high molecular weight protein produced by the LS174T colon carcinoma cell line. The use of shorter peptides indicated that the minimum peptide epitopes for these MAbs were different. Mab 3A2 reacted with amino acids 5-19 of the MUC2 VNTR by inhibition ELISA but not by direct ELISA, while 4FI reacted with this peptide in both assays. Furthermore, 4FI reacted in direct ELISA when a larger (29 amino acid) MUC2-derived peptide was coated onto the assay plate by incubating in carbonate buffer or by drying the peptide onto the assay plate, while 3A2 only reacted when this peptide was coated in carbonate buffer. The different specificity of the MAbs was also illustrated by the reactivity of 4F1 but not 3A2 with partially deglycosylated cystic fibrosis mucin.Immunohistochemical analysis with these MAbs revealed a strong reactivity with lung, gastric and colon tumours relative to normal tissue, with some breast and ovarian tumours also reacting. Both MAbs stained some normal goblet cells in the perinuclear region but not the mucin droplet or secreted mucin, indicating a reaction with immature (poorly glycosylated) mucin in the endoplasmic reticulum and/or golgi, but not with mature (fully glycosylated) mucin. In contrast, tumours showed strong diffuse cytoplasmic staining. 4F1 also showed weak apical cytoplasmic staining in some goblet cells and stained some tumours which showed no reactivity with 3A2.These antibodies should prove useful in the study of MUC2 structure and function, and in the diagnosis of some tumours.

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Section: Mucinsmentioning

confidence: 99%

Section: Elisamentioning

confidence: 99%

mentioning

confidence: 99%

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Monoclonal antibodies reacting with the MUC2 mucin core protein

Pl¹,

McGuckin²,

Birrell³

et al. 1993

Br J Cancer

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“…The plate was incubated for 3 h at RT (BC2 or CCP58) or overnight at 4 °C (3E1.2) before washing as described above and adding 50 pl/well of antimouse Ig-HRP at 1/1,000 in PBS-Tween for 1 h at RT. After washing the plate, peroxidase activity was visualized with ABTS substrate, and inhibition was calculated as described [26],…”

Section: Dual-determinant Elisa (Dde)mentioning

confidence: 99%

“…The assigned numbers for cell culture supernatants are as in table 1. Inhibition was calcu lated as described [26]. CCP58 are exposed on the MUC2 VNTR.…”

Section: Detection O F Muc1 and Muc2 In Culture Supernatantsmentioning

confidence: 99%

Expression of MUC1 and MUC2 Mucins by Human Tumor Cell Lines

Devine¹,

Birrell²,

Whitehead³

et al. 1992

Tumor Biol

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The secretion and nature of mucins produced from a panel of recently available new gastric and colon carcinoma cell lines (LIM1839, LIM1215, LIM1863, LIM1899, LIM2099, LIM2405, LIM2408, LIM2412, LIM2463), as well as other colon (LS174T, HT29, HT29-SB, COL0533, COLO206), breast (T47D, MCF-7, BT20, ZR75–1) and ovarian (COLO316) tumor cell lines, was investigated. ELISA and Western blotting of the culture supernatants with novel anti-MUC1 and anti-MUC2 monoclonal antibodies (MAbs) specific for mucin core proteins showed their secretion by most of these cell lines. In addition, mucins produced by these cell lines expressed the tumor-associated carbohydrate detected by MAb 3E1.2 (glycolylsialyl-Tn, mammary serum antigen or MSA) and the Tn or T antigens reactive with lectin SSA-M. SSA-M detected MUC1 or MUC2 captured by MAbs BC2 or CCP58, while 3E1.2 only detected MUC1-associated carbohydrate, indicating that the MAb may react with a conforma-tionally dependent epitope, or that the sialyl/glycolyl-transfer-ases involved in MSA production may be sequence specific. In addition, the BC2/SSA-M and CCP58/SSA-M assays detected mucins in some samples which were not detected by BC2/BC2 or CCP58/CCP58 dual determinant assays, indicating that this format may be more appropriate for the detection of tumor-associated mucins in body fluids. These new cell lines and assays should be of use in the investigation of mucin core proteins, particularly LIM2463 and LIM1839 which express significant quantities of both MUC1 and MUC2.

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Expression of the tumor‐associated mucin MUCI in an ovarian tumor cell line

Stern

Palatsides

Kretser

et al. 1992

Intl Journal of Cancer

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Epithelial sialomucins constitute a family of high-molecular-weight glycoproteins associated with epithelial cell surfaces. Aberrant expression of these molecules has been observed in certain types of human epithelial tumors. Members of the MUC1 family of mucins isolated from different tissue types have been shown to differ in biochemical properties and in immunological reactivity. The polymerase chain reaction (PCR) has been used in a study of the MUC1 mRNA expressed in ovarian tumor cells. The intron/exon structure of the ovarian mucin gene has been examined, the nucleotide sequence of regions of the cDNA 5' and 3' to a central highly repetitive region of the molecule determined and genomic clones for this mucin from the ovarian tumor cell line COLO316 have been isolated and analyzed. The results are compared with the nucleotide sequence data obtained by others for MUC1 cDNA in breast and pancreatic cell lines. With a single nucleotide exception, the splicing pattern and nucleotide sequence obtained from MUC1 mRNA in the ovarian cell line is the same as that of the mRNA found in breast and pancreatic cell lines. However, it appears that the use of alternate splice acceptors for intron I in the ovarian cell line studied here is independent of the specific sequence variation thought to determine splicing of MUC1 in breast tumor cells.

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Monoclonal Antibodies Reactive with the Breast Carcinoma-Associated Mucin Core Protein Repeat Sequence Peptide Also Recognise the Ovarian Carcinoma-Associated Sebaceous Gland Antigen

Cited by 29 publications

References 14 publications

Monoclonal antibodies reacting with the MUC2 mucin core protein

Monoclonal antibodies reacting with the MUC2 mucin core protein

Expression of MUC1 and MUC2 Mucins by Human Tumor Cell Lines

Expression of the tumor‐associated mucin MUCI in an ovarian tumor cell line

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