Preparation and characterization of anti-human chromogranin A and chromogranin B (secretogranin I) monoclonal antibodies

• Published in: Molecular and cellular probes Vol. 3; no. 1; pp. 87 - 101
• Main Authors: Pelagi, Micaela, Bisiani, Chiara, Gini, Ambrogio, Bonardi, Maria Antonietta, Rosa, Patrizia, Marè, Paolo, Viale, Giovanna, Cozzi, Maria Grazia, Salvadore, Maurizio, Zanini, Antonia, Siccardi, Antonio G., Buffa, Roberto
• Format: Journal Article
• Language: English
• Published: Kidlington Elsevier Ltd 01.03.1989; Elsevier
• Subjects: Antibodies, immunoglobulins; Antibodies, Monoclonal - immunology; Antibody Specificity; Biological and medical sciences; Blotting, Western; chromaffin granules; Chromaffin Granules - immunology; Chromaffin System - immunology; Chromogranin A; Chromogranin B; chromogranins; Chromogranins - immunology; Electrophoresis, Gel, Two-Dimensional; Fundamental and applied biological sciences. Psychology; Fundamental immunology; Humans; immunocytochemistry; Immunoenzyme Techniques; Molecular immunology; Monoclonal antibodies; Nerve Tissue Proteins - immunology; pheochromocytoma; two-dimensional immunoblotting; chromaffin granules; pheochromocytoma; immunocytochemistry; chromogranins; two-dimensional immunoblotting; Human; Immunization; Immunocytochemistry; Rodentia; Monoclonal antibody; Immunological method; Characterization; Proteins; Vertebrata; Mammalia; Chromaffin granule; Mouse; Secretory protein; Immunoblotting assay; Tumor
• ISSN: 0890-8508; 1096-1194
• DOI: 10.1016/0890-8508(89)90039-X

Chromogranin A, chromogranin B/secretogranin I and chromogranin C/secretogranin II are acidic sulphated and phosphorylated secretory proteins present in a large number of endocrine and neuronal tissues. It has been suggested that these proteins may be useful immunohistochemical markers for human tumours of endocrine origin and their measurement in plasma has been proposed as a diagnostic tool in patients with these tumours. In order to obtain anti-human chromogranins/secretogranins antibodies for clinical applications, we immunized mice with whole chromaffin granules isolated from human pheochromocytoma. The immune sera analysed by two-dimensional immunoblotting were found to recognize chromogranins/secretogranins and other unidentified proteins and to react in immunocytochemistry with pheochromocytoma as well as with a number of endocrine cells of different types. Hybridoma supernatants obtained from the splenocytes of a hyperimmune mouse, screened with an enzyme-linked immunosorbent assay, were analysed by both immunocytochemistry and two-dimensional immunoblotting. By using this experimental approach we were able to identify several monoclonal antibodies against human chromaffin granule components. In particular, we have characterized one anti-human chromogranin A and one anti-human chromogranin B/ secretogranin I monoclonal antibody which showed a very specific pattern both in immunocytochemistry and in two-dimensional immunoblotting.