BLOOD Glucose measurement

Sample type
Glucose is measured in whole blood or serum (i.e., plasma). Historically, blood glucose values were given in terms of whole blood, but most laboratories now measure and report the serum glucose levels. Because red blood cells (erythrocytes) have a higher concentration of protein (e.g., hemoglobin) than serum, serum has a higher water content and consequently more dissolved glucose than does whole blood. To convert from whole-blood glucose, multiplication by 1.15 has been shown to generally give the serum/plasma level.
Collection of blood in clot tubes for serum chemistry analysis permits the metabolism of glucose in the sample by blood cells until separated by centrifugation. Red blood cells, for instance, do not require insulin to intake glucose from the blood. Higher than normal amounts of white or red blood cell counts can lead to excessive glycolysis in the sample with substantial reduction of glucose level if the sample is not processed quickly. Ambient temperature at which the blood sample is kept prior to centrifuging and separation of plasma/serum also affects glucose levels. At refrigerator temperatures, glucose remains relatively stable for several hours in a blood sample. At room temperature (25 °C), a loss of 7-10 mg/dL (or 0.4 mmol/L) of total glucose per hour should be expected in whole blood samples. Loss of glucose under these conditions can be prevented by using Fluoride tubes (i.e., gray-top) since fluoride inhibits glycolysis. However, these should only be used when blood will be transported from one hospital laboratory to another for glucose measurement. Red-top serum separator tubes also preserve glucose in samples after being centrifuged isolating the serum from cells.
Particular care should be given to drawing blood samples from the arm opposite the one in which an intravenous line is inserted, to prevent contamination of the sample with intravenous fluids. Alternatively, blood can be drawn from the same arm with an IV line after the IV has been turned off for at least 5 minutes, and the arm elevated to drain infused fluids away from the vein. Inattention can lead to large errors, since as little as 10% contamination with 5% dextrose (D5W) will elevate glucose in a sample by 500 mg/dl or more. Remember that the actual concentration of glucose in blood is very low, even in the hyperglycemic.
Arterial, capillary and venous blood have comparable glucose levels in a fasting individual. After meals venous levels are somewhat lower than capillary or arterial blood; a common estimate is about 10%.
Measurement techniques
Two major methods have been used to measure glucose. The first, still in use in some places, is a chemical method exploiting the nonspecific reducing property of glucose in a reaction with an indicator substance that changes color when reduced. Since other blood compounds also have reducing properties (e.g., urea, which can be abnormally high in uremic patients), this technique can produce erroneous readings in some situations (5 to 15 mg/dl has been reported). The more recent technique, using enzymes specific to glucose, are less susceptible to this kind of error. The two most common employed enzymes are glucose oxidase and hexokinase.
In either case, the chemical system is commonly contained on a test strip, to which a blood sample is applied, and which is then inserted into the meter for reading. Test strip shapes and their exact chemical composition vary between meter systems and cannot be interchanged. Formerly, some test strips were read (after timing and wiping away the blood sample) by visual comparison against a color chart printed on the vial label. Strips of this type are still used for urine glucose readings, but for blood glucose levels they are obsolete. Their error rates were, in any case, much higher.
Urine glucose readings, however taken, are much less useful. In properly functioning kidneys, glucose does not appear in urine until the renal threshold for glucose has been exceeded. This is substantially above any normal glucose level, and so is evidence of an existing severe hyperglycemic condition. However, urine is stored in the bladder and so any glucose in it might have been produced at any time since the last time the bladder was emptied. Since metabolic conditions change rapidly, as a result of any of several factors, this is delayed news and gives no warning of a developing condition. Blood glucose monitoring is far preferable, both clinically and for home monitoring by patients.
I. CHEMICAL METHODS
A. Oxidation-Reduction Reaction
1. Alkaline Copper Reduction
Folin Wu Method
Blue end-product
Benedict's method
Modification of Folin wu for Qualitative Urine Glucose
Nelson Somogyi Method
Blue end-product
Neocuproine Method
*
Yellow-orange color Neocuproine
Shaeffer Hartmann Somogyi
Uses the principle of Iodine reaction with Cuprous byproduct.
Excess I2 is then titrated with thiosulfate.
2. Alkaline Ferricyanide Reduction
Hagedorn Jensen
Colorless end product; other reducing substances interfere with reaction
B. Condensation
Ortho-toluidine Method
Uses aromatic amines and hot acetic acid
Forms Glycosylamine and Schiff's base which is emerald green in color
This is the most specific method, but the reagent used is toxic
Anthrone (Phenols) Method
Forms hydroxymethyl furfural in hot acetic acid
II. ENZYMATIC METHODS
A. Glucose Oxidase
Saifer–Gerstenfeld Method
Inhibited by reducing substances like BUA, Bilirubin, Glutathione, Ascorbic Acid
Trinder Method
uses 4-aminophenazone oxidatively coupled with Phenol
Subject to less interference by increases serum levels of Creatinine, Uric Acid or Hemoglobin
Inhibited by Catalase
Kodak Ektachem
A Dry Chemistry Method
Uses Reflectance Spectrophotometry to measure the intensity of color through a lower transparent film
Glucometer
Home monitoring blood glucose assay method
Uses a strip impregnated with a Glucose Oxidase reagent
B. Hexokinase
NADP as cofactor
NADPH (reduced product) is measured in 340 nm
More specific than Glucose Oxidase method due to G-6PO_4, which inhibits interfering substances except when sample is hemolyzed
Blood glucose laboratory tests
fasting blood sugar (i.e., glucose) test (FBS)
urine glucose test
two-hr postprandial blood sugar test (2-h PPBS)
oral glucose tolerance test (OGTT)
intravenous glucose tolerance test (IVGTT)
glycosylated hemoglobin (HbA1C)
self-monitoring of glucose level via patient testing
Clinical correlation
The fasting blood glucose level, which is measured after a fast of 8 hours, is the most commonly used indication of overall glucose homeostasis, largely because disturbing events such as food intake are avoided. Conditions affecting glucose levels are shown in the table below. Abnormalities in these test results are due to problems in the multiple control mechanism of glucose regulation.
The metabolic response to a carbohydrate challenge is conveniently assessed by a postprandial glucose level drawn 2 hours after a meal or a glucose load. In addition, the glucose tolerance test, consisting of several timed measurements after a standardized amount of oral glucose intake, is used to aid in the diagnosis of diabetes. It is regarded as the gold standard of clinical tests of the insulin / glucose control system, but is difficult to administer, requiring much time and repeated blood tests. In comparison, the fasting blood glucose level is a much poorer screening test because of the high variability of the experimental conditions such as the carbohydrate content of the last meal and the energy expenditure between the last meal and the measurement. Actually, many people with prediabetes or diabetes can have a fasting blood glucose below the prediabetic/diabetic threshold if their last meal happened to be low in carbohydrate and they burnt all the related glucose in their blood stream before taking the test. Note that food commonly includes carbohydrates which don't participate in the metabolic control system; simple sugars such as fructose, many of the disaccarhides (which either contain simple sugars other than glucose or cannot be digested by humans) and the more complex sugars which also cannot be digested by humans. And there are carbohydrates which are not digested even with the assistance of gut bacteria; several of the fibres (soluble or insoluble) are chemically carbohydrates. Food also commonly contains components which affect glucose (and other sugar's) digestion; fat, for example slows down digestive processing, even for such easily handled food constituents as starch. Avoiding the effects of food on blood glucose measurement is important for reliable results since those effects are so variable.
Error rates for blood glucose measurements systems vary, depending on laboratories, and on the methods used. Colorimetry techniques can be biased by color changes in test strips (from airborne or finger borne contamination, perhaps) or interference (e.g., tinting contaminants) with light source or the light sensor. Electrical techniques are less susceptible to these errors, though not to others. In home use, the most important issue is not accuracy, but trend. Thus if your meter / test strip system is consistently wrong by 10%, there will be little consequence, as long as changes (e.g., due to exercise or medication adjustments) are properly tracked. In the US, home use blood test meters must be approved by the Federal Food and Drug Administration before they can be sold.
Finally, there are several influences on blood glucose level aside from food intake. Infection, for instance, tends to change blood glucose levels, as does stress either physical or psychological. Exercise, especially if prolonged or long after the most recent meal, will have an effect as well. In the normal person, maintenance of blood glucose at near constant levels will nevertheless be quite effective.
Causes of Abnormal Glucose Levels
Persistent Hyperglycemia
Transient Hyperglycemia
Persistent Hypoglycemia
Transient Hypoglycemia
Reference Range, FBG: 70–110 mg/dl
Diabetes Mellitus
Pheochromocytoma
Insulinoma
Acute Alcohol Ingestion
Adrenal cortical hyperactivity Cushing's Syndrome
Severe Liver Disease
Adrenal cortical insufficiency Addison's Disease
Drugs: salicylates, antituberculosis agents
Hyperthyroidism
Acute stress reaction
Hypopituitarism
Severe Liver disease
Acromegaly
Shock
Galactosemia
Several Glycogen storage diseases
Obesity
Convulsions
Ectopic Insulin production from tumors
Hereditary fructose intolerance