Insulin testing: Preparation, Reference Range and Understanding the Result

Insulin testing is used to assist in identifying causes of hypoglycemia, especially upon signs and symptoms of hypoglycemia which could be neurohypoglycopenic and autonomic.

Updated: October 11, 2019

Insulin testing is used to assist in identifying causes of hypoglycemia, especially upon signs and symptoms of hypoglycemia which could be neurohypoglycopenic and autonomic.
Hypoglycemia is a condition in which plasma glucose levels is less then 55 mg/dL. A 72-hour fasting test is performed in this case. Insulin testing is also used to diagnose early type 2 diabetes, in which there is a relatively increased production of insulin with a concurrent increase in blood glucose levels.
In addition, insulin testing is used to help differentiate type 1 and type 2 diabetes.
Insulin is an anabolic hormone which is responsible for promoting glucose uptake, glycogenesis, lipogenesis, and protein synthesis of skeletal muscle and fat tissue through the tyrosine kinase receptor pathway. It also helps in the regulation of plasma glucose homeostasis, as it counteracts glucagon and other catabolic hormones such as, epinephrine, glucocorticoid, and growth hormone. Insulin secretion is induced by elevated plasma glucose levels. Insulinoma is an example of hypersecretory state, in which insulin is secreted at a high rate independent from the plasma glucose level. Here even in hypoglycemic state, high insulin levels are found. Oral administration of glucose is more effective in increasing insulin secretion than intravenous glucose .

Preparation of Insulin testing:

A 72-hour fasting test is used to identify causes of hypoglycemia. The patient is instructed to fast, and plasma glucose, insulin, proinsulin, and C-peptide levels are measured every 6 hours until the plasma glucose level is less than 65 mg/dL. Once the plasma glucose level is less than 65 mg/dL, the testing frequency is increased to every 1 to 2 hours. When plasma glucose levels are less than 45 mg/dL accompanied by signs and symptoms of hypoglycemia, a blood sample is collected and tested for glucose, insulin, proinsulin, C-peptide, beta-hydroxybutyrate, and sulfonylurea levels. The response of the blood glucose level is measured after giving 1 mg of intravenous glucagon to the patient.

Reference Range:

Insulin level should be less then 25 mIU/L when done on fasting. After 30 minutes of glucose administration it should be 30 to 230 mIU/L. 1 hour after glucose administration it should be 18 to 276 mIU/L and 16 to 166 mIU/ after 2 hour. 3 hours after glucose administration it should return to normal range that is less then 25 mIU/L.
Looking in to the result of insulin test and C-peptide test it is easier to diagnose certain medical condition. High insulin and C-peptide levels indicate a condition called insulinoma, insulin secretagogue and congenital hyperinsulinism in which mutation in insulin-secreting gene happens.
High insulin levels and low C-peptide levels indicates excessive insulin administration. An extremely elevated insulin level and high insulin–to–C-peptide ratio is an indication of autoimmunity to insulin or insulin receptor. This is common in patients receiving insulin or those who have autoimmune diseases such as systemic lupus erythematosus (SLE) or Hashimoto thyroiditis. The cause is due to dissociation of postprandial insulin that is bound antobodies after 1 hour. Low insulin and C-peptide levels and high growth factor 2 level indicates non–beta cell tumors.

Understanding the Result:

Conditions associated with elevated insulin levels:

Conditions associated with increased insulin resistance include:

  • Obesity
  • Intake of steroid 
  • Acromegaly
  • Cushing syndrome
  • Insulin receptor mutation
  • Type 2 diabetes in early stage

In all these cases beta cell compensates via hypersecretion of insulin.

Conditions associated with increased insulin secretion include the following:

  • Insulinoma (insulin or proinsulin secreting tumors)
  • Administration of insulin secretagogues

Conditions associated with decreased insulin excretion include:

  • Severe liver disease
  • Severe heart failure

Conditions associated with decreased insulin levels:

Conditions associated with beta-cell destruction include the following:

  • Post pancreatectomy
  • Chronic pancreatitis
  • Type 1 diabetes 
  • Autoimmune destruction
  • Beta cells fail to secrete insulin for maintaining the blood glucose level, leading to insulin resistance and genetic defect in late stage of type 2 diabetes.

Factors affecting the test:

Insulin levels may be falsely elevated by the following:

  • Amino acid such as leucine, arginine, and lysine
  • Steroid
  • Insulin secretagogue such as sulfonylurea and glinide
  • Estrogen
  • Beta2 agonist

Insulin levels may be falsely decreased by the following:

  • Acarbose, metformin, octreotide, and beta-blocker
  • Hemolysis (insulin-degrading enzyme in red blood cell released)
  • Hemodialysis


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