1. Lab (7): Diabetic profile
KAU-Faculty of Science- Biochemistry department
Clinical biochemistry lab (Bioc 416) 2012
T.A Nouf Alshareef

2. Diabetes Mellitus:
It is a chronic disease due to disorder of carbohydrate metabolism, due to insulin deficiency results in hyperglycemia (increased blood glucose level) & glucourea (presence of glucose in urine).
Associated with several changes in metabolism; such as metabolism of proteins & fats.

3. Clinical Biochemical Findings in Diabetes:
Glucosuria.
Large volume of urine & increase urination frequency (Polyuria)
Polyphagia (eats more frequently).
Several metabolic changes

4. Metabolic changes in diabetes
Include increase in:
Fat catabolisim leads to increase in FFAs in blood & liver.
Acetyl.coA leads to increase formation of cholesterol & risk of atherosclerosis.
ketone bodies generation in blood and urine leads to acidosis.
catabolism of tissue protein due to energy requirement (because glucose can't uptake by cells) lead to weight loss and increase in level of amino acids in blood & more formation of urea by deamination of amino acid.

5. Types of diabetes: Type I diabetes mellitus (TIDM)
Type 2 diabetes mellitus (TIIDM)
Gestational diabetes mellitus (GDM)
Other "due to drugs or chemicals"

6. Diabetic profile:
- Is group of tests that are used to diagnose diabetes or its complications , it includes:
C-peptide.
Blood glucose (4 types: FBS, PPBS, RBS, OGGT)
HbA1C
Insulin
ICA (islent cell antibody) for type I
Ketones
Microalbuminurea.

7. 1- C-peptide:
This test is used to differentiate between type I and II diabetes
type I DM: low level of insulin and C-peptide
type II DM: normal or high level of C-peptide.
Advantages of measuring C-peptide than insulin:
It is better indicator of B-cell function than peripheral insulin.
Normal value: Fasting ng/ml
Human insulin and C-PEPTIDE originate as a single polypeptide chain known as proinsulin(M.Wt 9000) in the pancreatic _ cell. Proinsulin is cleaved proteolytically to form equimolar amounts of mature insulin and C-PEPTIDE that are released into the portal vein. So called because it connects the A and B chains of insulin in the proinsulin molecule. C-PEPTIDE is a single chain of 31 amino acid (Mol. Wt 30200).Unlike insulin has no known physiological function. Because C-PEPTIDE has a longer half-life than insulin (2-5 times longer), high concentrations of C-PEPTIDE persist in the peripheral circulation and these levels fluctuate less insulin. For these reasons, in plasma C-PEPTIDEe concentrations may reflect pancreatic insulin secretion more reliable than the level of insulin itself. C-PEPTIDE is cleaved from the body by the kidney. Urine concentrations of C-PEPTIDE are times higher than in plasma, unlike plasma insulin levels, which fluctuate in response to meals, measurement of the 24 hour urinary excretion of C-PEPTIDE provides a useful monitor of average cell insulin secretion2.C- PEPTIDE measurements are useful in insulinoma diagnosis, especially in patients treated with insulin. Elevated C-PEPTIDE levels are indicative of insulinoma. C-PEPTIDE measurements are useful in the need for progression to insulin therapy in non-insulin dependent diabetics (NIDDM). C-PEPTIDE measurements are useful as a marker for residual pancreatic tissue after pancreatectomy. It may also be used to monitor the progress of pancreas or islet cell transplantation. C-PEPTIDE measurements are useful in the diagnosis of hypoglycemia brought on by surreptitious insulin administration.

8. 2- Blood sugars: Types of blood glucose tests:
(1) Fasting blood sugar (FBS):
measures blood glucose after fasting for at least 8-12 hrs
It often is the first test done to check for diabetes.
patient with mild or borderline diabetes may present with normal FBG values.
If diabetes is suspected, GTT can confirm the diagnosis.
Normal levels:
60-110mg/dl

9. (2) Post-Prandial Blood Sugar (2-hour PPBS):
After the patient fasts for 12 hours, a meal is given which contains starch and sugar (approx. 100 gm).
Then after 2 hours blood is collected to measure glucose level.
home blood sugar test is the most common way to check 2-hour postprandial blood sugar levels.

10. (3) Random blood sugar (RBS)
measures blood glucose randomly at any time throughout the day without patient fasting.
it is useful because glucose levels in healthy people don’t vary widely throughout the day.
blood glucose levels that vary widely may indicate a problem.

11. (4) Oral glucose tolerance test (OGTT)
Glucose Tolerance is defined as the capacity of the body to tolerate an extra load of glucose or it measures the body's ability to use glucose.
it is series of blood glucose measurements taken after drink glucose liquid
It is considered as definitive diagnostic test for DM.
It is ordered to:
- confirm the diagnosis, in pre-diabetic - diagnose gestational diabetes (most commonly)
Recommended if mg/dl or (5.5 mmol/L-7.0 mmol/L).

12. Procedure:
Arrive FBS: After an overnight fasting of hrs
Drink: g dissolved in ml of water and given orally.
After drink: blood samples and urine are collected every 30min for 3hrs (1 hr, 1.5 hr , 2hr, 2.5hr, 3hr )
A curve between time and blood glucose concentration, is plotted.

13. Other types of OGTT:
Extended GTT :
glucose measured for 4-5 hrs after giving glucose to see how the curve behaves below the normal fasting glucose limits. Done in some conditions causing hypoglycaemia.
Cortisone Stressed GTT : Can be used for detecting latent DM.
Intrevenous GTT :
- is done if oral glucose is not tolerated or oral GTT curve is flat.
- In these cases 20% glucose as 0.5g glucose/Kg body weight.
- Usually peak occurs within 30 min after infusion and returns to normal after 90 min.

14. Interpretation: Normal Response :
FBS is normal. After 1 hr it will rise, returns to normal fasting level within 2 hours.
Diabetic curve :
FBS: 140mg/dl or 7.8 mmol/L. After 2 hr: 200mg/dl (11 mmol/L) or more. Glucosuria is usually seen
Impaired GTT:
- with 2hrs glucose level between 140mg/dl - 200mg/dl - it is not abnormal but must be followed up for DM. 

16. Renal Glycosuria:
Curve is normal due to lowered renal threshold one or more samples of urine contain glucose.
Lag storage/Alimentary Type:
FBS is normal. Due to rapid absorption, maximum level is found at 30 min (180mg/dl). glucosuria is seen hypoglycaemic levels may be reached at end of 2 hours. 
Flat curve of enchanced glucose tolerance:
FBS is normal. Throughout the test the level does not vary + 20mg%.

17. 3- HbA1c: HbA1C: is glucose bound to hemoglobin
Measures blood glucose conc. over a longer period of time
it indicates how well diabetes has been controlled in the 2-3 months before the test.
The A1C level is directly related to complications from diabetes (lower the A1C level lower risk for complications)
Type of sample: whole blood in EDTA tube
Normal Values:
Glycohemoglobin A1c:4.5%-5.7%
Total glycohemoglobin:5.3%-7.5%
It is Hb+A1C/100

18. Expected range of Hb A1c:
Sugar: % to 7.0%
Sugar: % to 8.0%
Sugar: % to 14.0%

19. Hypoglycaemia : When blood glucose falls below 60 mg/dl. Causes:
Most commonly seen in overdose of insulin in treatment of DM.
Hypothroidism.
Insulin secreting tumours of pancrease – rare.
Hypoadrenahsm (Addison's disease)
Hypopitruitism.
Severe exercise.
Starvation.

20. Experiment: Measuring glucose level
Principle:
Glucose + H2O + O Gluconic acid + H2O2
2H2O2 + 4 aminoantipyrine + PHBS Quinoneimine dye + H2O
Red color
GOD
POD

21. Kit components Glucose Oxidase Reagent : mixture of:
glucose oxidase + peroxidase+ aminoantipyrine+ buffer
Glucose standard Reagent :
conc. 100mg/dl or 5.55 mmol/L

22. Glucose oxidase reagent
Procedure
Prepare the reaction as the following:
Mix, incubates at 37oC for 10min
Read abs at 510nm
Sample
Standard
Reagent blank
1ml
1 ml
Glucose oxidase reagent
0.01ml or
10 ml -
Sample (serum)
Glucose standard

23. Glucose conc. = Abs. sample X conc. standard
Calculations:
Glucose conc. = Abs. sample X conc. standard
Abs. standard ..