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Publications (3)14.61 Total impact

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    ABSTRACT: Two synthetic peptides from the predicted sequence of the human c-erbB-2 protein were synthesized and used to raise antisera in rabbits. The sequences chosen were identical to those in the homologous rat c-neu protein. The antibodies produced reacted with the immunizing peptides in ELISA but showed little or no cross-reaction with the partly homologous peptides found in equivalent positions in the human EGF receptor. Both antipeptide antibodies, and a monoclonal antibody (MAb) specific for the rat neu protein, immunoprecipitated a 185-kDa protein from 35S-methionine-labelled lysates prepared from a rat cell line known to express high levels of the c-neu protein. The antipeptide antibodies also recognized a protein of the same size in Western blots. In addition, both antipeptide antibodies immunoprecipitated a 190-kDa protein from labelled cell lysates prepared from human and monkey cells. Antibodies to one of the peptides, which showed no detectable cross-reaction with human, rat or monkey EGF receptor, were used to examine the expression of the c-erbB-2 protein on a variety of cultured cell types. Eleven transformed, I non-established and 2 immortalized cell types were examined by immunoprecipitation for their level of expression of the c-erbB-2 protein and of the EGF receptor. The numbers of EGF receptors varied widely between different cell lines, whereas the level of the c-erbB-2 protein, which was found on all of the cell types examined, was more constant. The number of c-erbB-2 molecules present was estimated by autoradiography to be about 100,000 per cell. The antibodies were then used to examine the location and level of expression of the human c-erbB-2 and rat c-neu proteins in normal tissues. Immunohistochemical staining showed that the c-erbB-2 protein was highly expressed in rat kidney proximal tubules and loop of Henle. The c-erbB-2 protein was also present on normal human epithelial cells but in some cases with a different distribution to that of the EGF receptor.
    International Journal of Cancer 09/1987; 40(2):246-54. · 5.01 Impact Factor
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    ABSTRACT: Two synthetic peptides from the predicted sequence of the human c-erbB-2 protein were synthesized and used to raise antisera in rabbits. The sequences chosen were identical to those in the homologous rat c-neu protein. The antibodies produced reacted with the immunizing peptides in ELISA but showed little or no cross-reaction with the partly homologous peptides found in equivalent positions in the human EGF receptor. Both antipeptide antibodies, and a monoclonal antibody (MAb) specific for the rat new protein, immuno-precipitated a 185-kDa protein from 35S-methionine-labelled lysates prepared from a rat cell line known to express high levels of the c-neu protein. The antipeptide antibodies also recognized a protein of the same size in Western blots. In addition, both antipeptide antibodies immunoprecipitated a 190-kDa protein from labelled cell lysates prepared from human and monkey cells. Antibodies to one of the peptides, which showed no detectable cross-reaction with human, rat or monkey EGF receptor, were used to examine the expression of the c-erbB-2 protein on a variety of cultured cell types. Eleven transformed, I non-established and 2 immortalized cell types were examined by immunoprecipitation for their level of expression of the c-erbB-2 protein and of the EGF receptor. The numbers of EGF receptors varied widely between different cell lines, whereas the level of the c-erbB-2 protein, which was found on all of the cell types examined, was more constant. The number of c-erbB-2 molecules present was estimated by autoradiography to be about 100,000 per cell. The antibodies were then used to examine the location and level of expression of the human c-erbB-2 and rat c-neu proteins in normal tissues. Immunohistochemical staining showed that the c-erbB-2 protein was highly expressed in rat kidney proximal tubules and loop of Henle. The c-enbB-2 protein was also present on normal human epithelial cells but in some cases with a different distribution to that of the EGF receptor.
    International Journal of Cancer 08/1987; 40(2):246 - 254. DOI:10.1002/ijc.2910400221 · 5.01 Impact Factor
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    ABSTRACT: A431 cells have been used as an immunogen for generating monoclonal antibodies against the epidermal growth factor (EGF) receptor. Two immunoglobulin M and eight immunoglobulin G3 anti-EGF receptor antibodies were cloned. All ten antibodies immunoprecipitated biosynthetically labeled mature A431 cell EGF receptor and were able to recognize the receptor in Western blotting. However, none of the antibodies immunoprecipitated precursor polypeptides of the A431 cell EGF receptor, neither did they recognize EGF receptors from human foreskin fibroblasts, human placenta, nor a human-mouse hybrid cell expressing EGF receptor. The antibodies were found to bind to glycolipids from A431 cells and it was shown that the determinant involved was the blood group A antigen. It appears that this determinant is present on both the EGF receptor and glycolipids of A431 cells but is not expressed on EGF receptors from other human cells tested. One of the monoclonal antibodies raised was used for immunoaffinity purification of the EGF receptor. The procedure took advantage of the carbohydrate nature of the antigenic determinant by employing sugar-specific elution. The mild conditions permitted the purification of A431 cell EGF receptor (70-80% pure) that possessed an intrinsic EGF-stimulated tyrosine kinase activity with a specific activity of about 20 nmol/min/mg.
    Journal of Biological Chemistry 09/1984; 259(15):9906-12. · 4.60 Impact Factor