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Diabetes is a common life-long health condition. There are 2.8 million people diagnosed with diabetes in the UK and an estimated 850,000 people who have the condition but do not know it.
Diabetes is a condition where the amount of glucose in your blood is too high because the body cannot use it properly. This is because your pancreas does not produce any insulin, or not enough, to help glucose enter your body's cells - or the insulin that is produced does not work properly (known as insulin resistance). To assess a patient with diabetes many blood tests need to be done to measure the plasma glucose level, such as (RBS), (FBS), and (OGTT).
Random blood glucose level (RBS): A sample of blood taken at any time can be a useful test if diabetes is suspected. A level of ≥ 11.1 mmol/L in the blood sample indicates that you have diabetes.
Oral glucose tolerance test (OGTT): is a medical test, which is used to measure the level of glucose in the blood and afterward to determine how fast it is cleared from the blood. This test is usually used to examine diabetes, insulin resistance, and reactive hypoglycaemia and sometimes carbohydrate metabolism. An oral glucose tolerance test (OGTT); is standard dose of glucose is given by mouth and blood levels are checked two hours later.
Glucose dehydrogenase method: is another enzymic method for glucose determination.By the action of glucose dehydrogenase enzyme (GDH) with NAD, D-glucose and NAD are transformed to D-gluconolactone and NADH. The change in NADH absorbance is measured in the UV region (334, 340 or 365 nm), because the amount of the reduced coenzyme formed is proportional to the concentration of glucose.
Table (1), Reference ranges for plasma glucose (OGTT) are:
< 7.8 mmol/L
< 7.0 mmol/L
≥ 7.8 mmol/L and
< 11.1 mmol/L
≥ 7.8 mmol/L
≥ 11.1 mmol/L
The Aim of this laboratory test is to practise and measure the plasma glucose level using the OGTT test, and to interpret the results according to the standard measures
Apparatus: Spectrophotometer, Pipettes, (24) LP4 tubes microliter, Gilson, Distell Water; (4) Blood, samples of Patient, Gloves, OGTT, Laboratory coat, Beaker, Paper towels.
Method: in the given protocol there are series of glucose standard concentration (240, 180, 160, 120, 60, and 0mg/dl) and amount of distilled water to prepare 500µL of water for each concentration. Using the formula C1xV1=C2xV2 to find out the volume (1) for each sample; as C1 was already known as 300mg/dl, C2 represents the series of glucose standards, while (V2) is 500 µL. Then the (V1) of each concentration was duplicated using Gilson to prepare two tubes containing 200 µL; so 10 tubes were collected in total. Having completed this step, a line graph plotted to find out the equation (y =0.0005X + 0.0038), which required measuring the plasma glucose concentration of each sample in mmol/L. Three patients fasted for 12 hours (A,B and C), the first blood samples aspirated directly on fasting , after that each patient took OGTT and blood samples collected 2 hours later. Then, each sample divided into two separated tubes with 200 µL in each one and labelled into (A0, B0, B2, C0, C2, and D), However, Patient D blood sample was collected directly without fasting and GOTT.
Overall,14 LP4 tubules were prepared with 200 µL in each one except the sample A2 was diluted 1+2 before use with distilled water. Finally, 2 ml of glucose oxidase peroxide (GOD/HRP) enzyme added to the 24 tubes. It had been recommended to leave the tubules in rest for 20 minutes to allow the colure to be ready and stabled. Finally, all the tubes were measured in the spectrophotometer machine to obtain their absorbance. .
Result: According to the figure (1), the five standard concentrations from 0 mg/dl to 240mg/dl were used to measure the mean absorbance, to find out the concentrations of plasma glucose by using the equation (y= 0.0005x +0.0038), (y) represents the mean absorbance of the samples and (x) represents the concentration of the samples.
An appropriate tube used as a reagent blank measured the absorbance of each at 575nm. Regarding to the table (2); the glucose concentration in sample (A), represents the highest level in both fasting and after 2-hour measurement, 9.0 mmol/L, and 16.9 mmol/L, respectively. Followed by patient (B); with 7.0mml/L in B0, and 11.3 mmol/L in B2 sample.
However, in patient (C), the reading was nearly identical in both C0 and C2 between 7.6 and 7.9 mmol/L. In patient (D) the measure was 6.7 mmol/L, the sample was measured as a random blood glucose test.
Blood concentration. in mmol/L
* The absorbance 1 and 2 has been adjusted to the blank 0.708
Discussion: Comparing the blood glucose results in table (2) with the reference range in table (1); they were located between impaired glucose tolerance (IGT) and diabetic cases. In patient A, the fasting measure was higher than 7.8 mmol/L, 2hours later the test was 16.9mmol/L; having got both tests high, it confirmed the presence of diabetes in this patient. However, in patient B, the fasting measure was 7.0mmol/L, which means it is an impaired glucose tolerance case; however, this result was strongly confirmed the diagnosis of diabetes, as the reading of blood glucose level was 11.3 mmol/L. Therefore, to reach the correct diagnosis, many physicians would prefer to repeat the test if there is any suspicion in order to commence the convenient management. For instance, Diabetic patients need to stay on their medication for years, and the doctors recommend regular blood tests for their patients to measure the response of treatment, at the same time these tests can reduce incidence of the complications of the disease during its progress.
Regarding to the patient C, we can see that both the fasting and 2 hours later results were located in IGT group, in medical terms this patient need to take care about her/his lifestyle and should follow a conservative dietary regime to avoid having diabetes at any time; as he/she is more susceptible to become a diabetic. However, in patient D, the random blood glucose level was 6.1mmol/L, it means that it is a normal level, clinically it does not rule out diabetes as many doctors recommend and adopt, for instance, the OGTT to confirm the diagnosis. Overall, the GOTT play a major role in medicine to diagnose the diabetes and other diseases such as Acromegaly.