C-peptide is the abbreviation for connecting peptide; it is a 31-amminoacid peptide. C-peptide of insulin is the C-terminal cleavage product produced during processing of the insulin prohormone to the mature insulin molecule. Proinsulin is cleaved when it is released from the pancreas into the blood - one C-peptide for each insulin molecule. C-Peptide is devoid of any biological activity but appears to be necessary to maintain the structural integrity of Insulin.

In-vitro determination of Insulin and C-Peptide level help in differential diagnosis of liver disease, acromegaly, Cusing syndrome, familial glucose intolerance, Insulinimia, renal failure, ingestion of accidental oral hypoglycaemic drugs or C-peptide induced factitious hypoglycaemia.

Newly diagnosed diabetes patient often get their C-peptide levels measured, to find if they have type 1 diabetes or type 2 diabetes. The pancreas of patients with type 1 diabetes is unable to produce insulin and they will therefore usually have a decreased level of C- peptide, while C-peptide levels in type 2 patients is normal or higher than normal. Measuring C-peptide in patients injecting insulin can help to determine how much of their own natural insulin these patients are still producing.  

C-peptide assays may be analytically more sensitive than insulin assays. Measurement of the C-peptide may be useful in evaluating endogenous insulin secretion in a variety of clinical conditions. Insulin and C-Peptide are secreted into portal circulation in equimolar concentrations; fasting levels of C-Peptide are 5 ? 10 fold higher than those of Insulin owing to the longer half-life of C-Peptide. The liver does not extract C-Peptide however; it is removed from the circulation by degradation in the kidneys with a fraction passing out unchanged in urine. Hence the urine C-Peptide levels correlate well with fasting C-Peptide levels in serum.

Intended Use

Immunoenzymatic colorimetric method for quantitative determination of C-Peptide in human serum.

Principle of the Assay

In this method, C-Peptide calibrators, patient specimens and/or controls containing the native antigen are first added to streptavidin coated wells. Biotinylated monoclonal and horseradish peroxidase (HRP) labelled antibodies are added and the reactants are mixed.

The different types of antibodies used have high affinity and specificity and are directed against distinct and different epitopes of C- Peptide. Reaction between the various C-Peptide antibodies and native C-Peptide occurs in the microwells without competition or steric hindrance forming a soluble sandwich complex.

Simultaneously, the complex is fixed to the well through the high affinity reaction of streptavidin and biotinylated antibody.

After equilibrium is attained, the antibody-bound fraction is separated from unbound antigen by aspiration. The native antigen concentration is directly proportional to the HRP activity in the antibody-bound fraction. The activity of the conjugated HRP is quantitated by reaction with TMB substrate to produce blue color. The reaction is terminated by adding stop solution which turns the blue color into yellow. The absorbance is measured on a plate reader.                                                                 

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