Sources/Clones
Antibodies to chromogranin A are available from Accurate (A3), Biogenesis (A11, LK2H10), Biogenex (A11, LK2H10), Camon (LK2H10), Cymbus Bioscience (LK2H10), Dako (DAK-A3, polyclonal), Diagnostic Biosystems (DAK-A3), Enzo (PHE5), Immunotech (LK2H10, C3420), Milab (CH), RDI (LK2H10), Novocastra (LK2H10), Medac, Sanbio (LK2H10), Saxon, Serotec (LK2H10, C3420) and Zymed.
Fixation/Preparation
The antibodies are immunoreactive in fixed, paraffin-embedded sections and frozen sections. HIER does not result in significant enhancement. Fixation in Bouin's or B5 fixative may improve immunogenicity. Proteolytic digestion does not improve immunostaining.
Background
The chromogranins are a family of soluble acidic proteins of about 68 kD. They are the major proteins in the peptide-containing dense core (neurosecretory) granules of neuroendocrine cells and sympathetic nerves. Ultrastructural examination has confirmed the localization of chromogranins to the matrix of neurosecretory granules of neuroendocrine cells. While having different molecular weights, the chromogranin subunits are neither identical nor entirely dissimilar and may differ in only two or three amino acid residues, with a minimum homology between any pair of polypeptides of about 33%. The chromogranins in neuroendocrine tissues display both quantitative and qualitative variability. They occur in the highest concentration in the following rank order: the adrenal medulla; anterior, intermediate and posterior pituitary; pancreatic islets; small intestine; thyroid C cells; and hypothalamus.
The antibody clone LK2H10 to chromogranin of 68 kD labels most normal neuroendocrine cells and their corresponding neoplasms. The LK2H10 clone was derived from human pheochromocytoma and exhibits crossreactivity with monkey and pig chromogranins (Wilson & Lloyd, 1984).
Chromogranins are thought to stabilize the soluble portion of neurosecretory granules by interaction with adenosine triphosphate and catecholamines and are released into the serum after splanchnic stimulation. They have multiple roles in the secretory process of hormones. Intracellularly, they are involved in targeting peptide hormones and neurotransmitters to granules of the regulated pathway by virtue of their ability to aggregate in the low-pH, high-calcium environment of the trans-Golgi network. Extracellular peptides formed as a result of proteolytic processing of chromogranins regulate hormone secretion. The synthesis of chromogranins is regulated by many different factors, including steroid hormones and agents that act through a variety of signaling pathways (Hendy et al, 1995).
Applications
The major applications of antibodies to the chromogranins are for the identification of neuroepithelial/neuroendocrine differentiation in normal and neoplastic tissues (Hirose et al, 1995; Blumenfeld et al, 1996), as well as the neural elements of the brain (Schiffer et al, 1995) and gut (Shen et al, 1994). Initial experience with clone LK2H10 to chromogranin A revealed less than 100% sensitivity for neuroendocrine cells, especially among those cells and tumors with low concentrations of neurosecretory granules and among tumors such as insulinomas, somatostatinomas, prolactinomas and corticotropin-and growth hormone-producing adenomas. However, the rate of positivity has improved with the use of more sensitive immunolabeling procedures.
Chromogranin is the most specific marker for neuroendocrine differentiation and corresponds to the neurosecretory granule, the hallmark of the neuroendocrine cell (Appendices 1.3, 1.19, 1.26). While it may be used with other neuroendocrine markers such as NSE and PGP9.5 to improve the diagnostic yield, chromogranin and synaptophysin are the most specific of all neuroendocrine markers.
Comments
As neurosecretory granules tend to be localized beneath the plasma membranes of neuroendocrine cells, their highest density is within the cytoplasmic processes characteristic of such cells. As such, staining for chromogranin often highlights the cytoplasmic processes often not visible in H&E stains. Aberrant immunoreactivity for chromogranin has been described in normal and neoplastic urothelium, particularly in the umbrella cells, attributed to reactivity with chromogranin-like proteins in the transitional cells (Mai et al, 1994).
References
•Blumenfeld W, Chandhoke DK, Sagerman P, Turi GK 1996 Neuroendocrine differentiation in gastric adenocarcinomas. An immunohistochemical study. Archives of Pathology and Laboratory Medicine 120: 478-481.
•Hendy GN, Bevan S, Mattei MG, Mouland AJ. Chromogranin A 1995 Clinical Investigative Medicine 18:47-65.
•Hirose T, Scheithauer BW, Lopes MB, et al 1995 Olfactory neuroblastoma. An immunohistochemical, ultrastructural and flow cytometric study. Cancer 76:4-19.
•Mai KT, Perkins DG, Parks W et al 1994 Unusual immunostaining pattern of chromogranin in normal urothelium and in transitional cell neoplasms. Acta Histochemia 96: 303-308.
•Schiffer D, Cordera S, Giordana MT et al 1995 Synaptic vesicle proteins, synaptophysin and chromogranin A in amyotropic lateral sclerosis. Journal of Neurological Science 129:68-74.
•Shen Z, Larsson LT, Malmfors G, et al 1994 Chromogranin A and B in neuronal elements in Hirschsprung's disease: an immunocytochemical and radioimmunoassay study. Journal of Pediatric Surgery 29: 1293-1301.
•Wilson BS, Lloyd RV 1984 Detection of chromogranin in neuroendocrine cells with a monoclonal antibody. American Journal of Pathology 115: 458-468.
Bibliografía
Manual of diagnostic antibodies for immunohistology / Anthony S.-Y. Leong, Kumarasen Cooper, F. Joel W.-M. Leong.
Antibodies to chromogranin A are available from Accurate (A3), Biogenesis (A11, LK2H10), Biogenex (A11, LK2H10), Camon (LK2H10), Cymbus Bioscience (LK2H10), Dako (DAK-A3, polyclonal), Diagnostic Biosystems (DAK-A3), Enzo (PHE5), Immunotech (LK2H10, C3420), Milab (CH), RDI (LK2H10), Novocastra (LK2H10), Medac, Sanbio (LK2H10), Saxon, Serotec (LK2H10, C3420) and Zymed.
Fixation/Preparation
The antibodies are immunoreactive in fixed, paraffin-embedded sections and frozen sections. HIER does not result in significant enhancement. Fixation in Bouin's or B5 fixative may improve immunogenicity. Proteolytic digestion does not improve immunostaining.
Background
The chromogranins are a family of soluble acidic proteins of about 68 kD. They are the major proteins in the peptide-containing dense core (neurosecretory) granules of neuroendocrine cells and sympathetic nerves. Ultrastructural examination has confirmed the localization of chromogranins to the matrix of neurosecretory granules of neuroendocrine cells. While having different molecular weights, the chromogranin subunits are neither identical nor entirely dissimilar and may differ in only two or three amino acid residues, with a minimum homology between any pair of polypeptides of about 33%. The chromogranins in neuroendocrine tissues display both quantitative and qualitative variability. They occur in the highest concentration in the following rank order: the adrenal medulla; anterior, intermediate and posterior pituitary; pancreatic islets; small intestine; thyroid C cells; and hypothalamus.
The antibody clone LK2H10 to chromogranin of 68 kD labels most normal neuroendocrine cells and their corresponding neoplasms. The LK2H10 clone was derived from human pheochromocytoma and exhibits crossreactivity with monkey and pig chromogranins (Wilson & Lloyd, 1984).
Chromogranins are thought to stabilize the soluble portion of neurosecretory granules by interaction with adenosine triphosphate and catecholamines and are released into the serum after splanchnic stimulation. They have multiple roles in the secretory process of hormones. Intracellularly, they are involved in targeting peptide hormones and neurotransmitters to granules of the regulated pathway by virtue of their ability to aggregate in the low-pH, high-calcium environment of the trans-Golgi network. Extracellular peptides formed as a result of proteolytic processing of chromogranins regulate hormone secretion. The synthesis of chromogranins is regulated by many different factors, including steroid hormones and agents that act through a variety of signaling pathways (Hendy et al, 1995).
Applications
The major applications of antibodies to the chromogranins are for the identification of neuroepithelial/neuroendocrine differentiation in normal and neoplastic tissues (Hirose et al, 1995; Blumenfeld et al, 1996), as well as the neural elements of the brain (Schiffer et al, 1995) and gut (Shen et al, 1994). Initial experience with clone LK2H10 to chromogranin A revealed less than 100% sensitivity for neuroendocrine cells, especially among those cells and tumors with low concentrations of neurosecretory granules and among tumors such as insulinomas, somatostatinomas, prolactinomas and corticotropin-and growth hormone-producing adenomas. However, the rate of positivity has improved with the use of more sensitive immunolabeling procedures.
Chromogranin is the most specific marker for neuroendocrine differentiation and corresponds to the neurosecretory granule, the hallmark of the neuroendocrine cell (Appendices 1.3, 1.19, 1.26). While it may be used with other neuroendocrine markers such as NSE and PGP9.5 to improve the diagnostic yield, chromogranin and synaptophysin are the most specific of all neuroendocrine markers.
Comments
As neurosecretory granules tend to be localized beneath the plasma membranes of neuroendocrine cells, their highest density is within the cytoplasmic processes characteristic of such cells. As such, staining for chromogranin often highlights the cytoplasmic processes often not visible in H&E stains. Aberrant immunoreactivity for chromogranin has been described in normal and neoplastic urothelium, particularly in the umbrella cells, attributed to reactivity with chromogranin-like proteins in the transitional cells (Mai et al, 1994).
References
•Blumenfeld W, Chandhoke DK, Sagerman P, Turi GK 1996 Neuroendocrine differentiation in gastric adenocarcinomas. An immunohistochemical study. Archives of Pathology and Laboratory Medicine 120: 478-481.
•Hendy GN, Bevan S, Mattei MG, Mouland AJ. Chromogranin A 1995 Clinical Investigative Medicine 18:47-65.
•Hirose T, Scheithauer BW, Lopes MB, et al 1995 Olfactory neuroblastoma. An immunohistochemical, ultrastructural and flow cytometric study. Cancer 76:4-19.
•Mai KT, Perkins DG, Parks W et al 1994 Unusual immunostaining pattern of chromogranin in normal urothelium and in transitional cell neoplasms. Acta Histochemia 96: 303-308.
•Schiffer D, Cordera S, Giordana MT et al 1995 Synaptic vesicle proteins, synaptophysin and chromogranin A in amyotropic lateral sclerosis. Journal of Neurological Science 129:68-74.
•Shen Z, Larsson LT, Malmfors G, et al 1994 Chromogranin A and B in neuronal elements in Hirschsprung's disease: an immunocytochemical and radioimmunoassay study. Journal of Pediatric Surgery 29: 1293-1301.
•Wilson BS, Lloyd RV 1984 Detection of chromogranin in neuroendocrine cells with a monoclonal antibody. American Journal of Pathology 115: 458-468.
Bibliografía
Manual of diagnostic antibodies for immunohistology / Anthony S.-Y. Leong, Kumarasen Cooper, F. Joel W.-M. Leong.
