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Synonyms = Insulin, IDDM , IDDM1 , IDDM2 , ILPR , IRDN , MODY10
Antibody type = Recombinant Rabbit monoclonal / IgG
Clone = HMV-363
Positive control = Pancreas: A strong C-Peptide staining should be seen in a large fraction of islet cells.
Negative control = Colon: C-Peptide staining must be absent in all cell types.
Cellular localization = Secreted
Reactivity = Human
Application = Immunohistochemistry
Dilution = 1:100 – 1:200
Intended Use = Research Use Only
Relevance of Antibody
C-Peptide is a surrogate marker for insulin producing cells.
C-peptide (connecting peptide) is a part of the proinsulin protein which is coded by the insulin gene at 11p15.5 and produced exclusively by the beta cells of the pancreatic islets. C-peptide is formed when the proinsulin is split into insulin and C-peptide. At that time equimolar quantities of insulin and C-peptide are released to the blood. C-peptide binds to the surface of several cell types (neuronal, endothelial, fibroblast and renal tubular) and can activate specific pathways. The clinical significance of C-peptide lies in its serological measurement as a parameter for endogenous insulin production (not influenced by exogenous insulin).
Staining Pattern in Normal Tissues
Images describing the C-peptide staining pattern in normal tissues obtained by the antibody HMV-363 are shown in our “Normal Tissue Gallery”.
|Respiratory system||Respiratory epithelium||Negative.|
|Gastrointestinal Tract||Salivary glands||Negative.|
|Pancreas||Strong cytoplasmic C-peptide immunostaining of the majority of islet cells. A faint staining of acinar cells surrounding pancreatic islets can be seen because of „contamination artifacts“.|
|Muscle/connective tissue||Heart muscle||Negative.|
|Bone marrow/lymphoid||Bone marrow||Negative.|
In normal tissues, C-peptide is only produced in islet cells of the pancreas
These findings are fully consistent with the RNA data described in the Human Protein Atlas (Tissue expression C-Peptide) which also describe insulin expression to be limited to the pancreas.
Positive control = Pancreas: A strong C-peptide staining should be seen in a large fraction of islet cells.
Negative control = Colon: C-peptide staining must be absent in all cell types.
Staining Pattern in Relevant Tumor Types
C-peptide expression is largely limited to insulinoma, an insulin producing neuroendocrine tumor of the pancreas. Extra-pancreatic insulinomas can occur but they are very rare.
The TCGA findings on C-Peptide RNA expression in different tumor categories have been summarized in the Human Protein Atlas.
Compatibility of Antibodies
No data available at the moment
IHC users have different preferences on how the stains should look like. Some prefer high staining intensity of the target stain and even accept some background. Others favor absolute specificity and lighter target stains. Factors that invariably lead to more intense staining include higher concentration of the antibody and visualization tools, longer incubation time, higher temperature during incubation, higher temperature and longer duration of the heat induced epitope retrieval (slide pretreatment). The impact of the pH during slide pretreatment has variable effects and depends on the antibody and the target protein.
All images and data shown here and in our image galleries are obtained by the manual protocol described below. Other protocols resulting in equivalent staining are described as well.
Freshly cut sections should be used (less than 10 days between cutting and staining). Heat-induced antigen retrieval for 5 minutes in an autoclave at 121°C in pH 7,8 Target Retrieval Solution buffer. Apply HMV-363 at a dilution of 1:150 at 37°C for 60 minutes. Visualization of bound antibody by the EnVision Kit (Dako, Agilent) according to the manufacturer’s directions.
Potential Research Applications
- To what extent ectopic production of C-peptide can occur in cancer has not been analyzed.
Evidence for Antibody Specificity in IHC
There are two ways how the specificity of antibodies can be documented for immunohistochemistry on formalin fixed tissues. These are: 1. Comparison with a second independent method for target expression measurement across many different tissue types (orthogonal strategy), and 2. Comparison with one or several independent antibodies for the same target and showing that all positive staining results are also seen with other antibodies for the same target (independent antibody strategy).
Orthogonal validation: For the antibody HMV-363, staining specificity for C-peptide is demonstrated by the complete concordance of the immunostaining data with data from three independent RNA screening studies, including the Human Protein Atlas (HPA) RNA-seq tissue dataset, the FANTOM5 project, and the Genotype-Tissue Expression (GTEx) project, which are all summarized in the Human Protein Atlas (Tissue expression C-Peptide). C-peptide immunostaining by HMV-363 is only seen in the pancreas (islet cells), the only organ for which insulin/C-peptide RNA expression had previously been documented.