Clinical Chemistry Review - Glucose, Iron, & Bilirubin Flashcards
conversion of glucose to pyruvic acid or lactic acid
glycolysis
production of glycogen from glucose
glycogenesis
production of glucose from glycogen
glycogenolysis
production of glucose from non-carbohydrate sources
gluconeogenesis
What is the end product of anaerobic glycolysis?
lactic acid
Which sugar accounts for nearly all blood sugar and is the body’s major source of cellular energy?
glucose
Name 5 hormones that regulate glucose levels and tell whether each raises or lowers the level.
- Insulin - lowers glucose
- Glucagon - increases glucose (has the greatest effect)
- Cortisol - increases glucose
- epinephrine - increases glucose
- growth hormone - increases glucose
What is the reference range for a fasting glucose in an adult?
70-99 mg/dL
Compare the normal blood glucose level for newborns and adults.
Glucose is lower in the newborn. The condition is known as physiologic hypoglycemia.
What are critical values for glucose?
<40 mg/dL and >600 mg/dL
What is glucosuria?
Glucose in the urine
What is glycosuria?
presence of any sugar in the urine
What is renal threshold?
The blood concentration of a substance that can be reabsorbed by the renal tubules.
What is the average renal threshold for glucose?
160-180 mg/dL
When the blood level exceeds this amount, glucose is excreted in the urine.
What is hyperglycemia?
High blood sugar.
List 4 causes of Type 1 diabetes mellitus.
- pancreatic beta cell destruction
- absence of insulin
- autoimmune - antibodies to insulin and islet cells
- genetic predisposition - HLA DR 3/4
List 2 causes of Type 2 diabetes mellitus.
- insulin resistance in peripheral tissue
- insulin secretory defect of beta cells
- associated with obesity
Compare Type 1 and Type 2 diabetes mellitus.
Type 1:
1. onset usually below age 40.
2. dependency on injected insulin
3. prone to ketoacidosis and diabetic complications
Type 2:
1. most common type
2. onset typically after age 40, but increasingly in teens
3. not dependent on exogenous insulin
4. not prone to ketoacidosis
Which tests are used to screen for diabetes mellitus?
- fasting plasma glucose
- 2-hour plasma glucose after a 75-g glucose load
- hemoglobin A1C
What are the criteria of the ADA for the diagnosis of diabetes mellitus?
Any of the following on 2 occasions:
1. random plasma glucose >=200 mg/dL
2. fasting plasma glucose >=126 mg/dL
3. 2-hour postload glucose >=200 mg/dL
OR
Hemoglobin A1C >=6.5% on more than one occasion, or in combination with results of one of the previously mentioned plasma glucose tests over the diagnostic threshold, is also diagnostic.
How is the 2-hour postload glucose performed?
Blood is drawn 2 hours after the patient ingests a 75-g glucose drink.
How is gestational diabetes diagnosed?
The woman is given a 50-g load of glucose at 24-28 weeks of gestation and blood is drawn 1 hour later.
A 1-hour plasma glucose >=140 mg/dL is abnormal and should be followed by a 3-hour oral glucose tolerance test (OGTT) using a 100-g glucose load after a fast of 8-14 hours.
Gestational diabetes is diagnosed by 2 or more of the following venous plasma glucose values:
1. fasting >=95 mg/dL
2. 1 hour >=180 mg/dL
3. 2 hour >=155 mg/dL
4. 3 hour >=140 mg/dL
What are the 2 most common methods for glucose determination?
- Glucose oxidase
- hexokinase
Describe the glucose oxidase method for glucose determination.
Glucose oxidase catalyzes the conversion of glucose to hydrogen peroxide and gluconic acid.
The second step of the reaction is a peroxidase reaction, which is much less specific than the glucose oxidase reaction.
Describe the hexokinase method for glucose determination.
Hexokinase catalyzes the phosphorylation of glucose to glucose-6-phosphate, which is then oxidized to G-6-PD in the presence of NADP+.
Why is the hexokinase method more accurate than the glucose oxidase method.
It is subject to fewer interfering factors.
A 60-year-old diabetic has a blood glucose of 210 mg/dL. His urine glucose is negative. Assuming there are no substances in his urine to inhibit the glucose reaction, what might account for the apparent discrepancy in these test results?
Diabetics may develop elevated renal thresholds.
That is why urine testing is not a good screening test for diabetes mellitus.
A diabetic patient who performs home glucose monitoring is sent to the local hospital lab for testing. The patient is suspicious of the quality of care he receives form his HMO, so immediately before going to the lab to have his blood drawn, he tests himself. His result is 128 mg/dL. The laboratory result is 150 mg/dL. Assuming that both tests were performed correctly and that controls were within the acceptable range, what might account for the discrepancy in values?
Home testing uses capillary whole blood; laboratory methods use venous plasma or serum.
Whole blood glucose is approximately 11% lower than plasma glucose.
In addition, different methodologies are used.
A specimen for a fasting glucose is drawn in a red-top tube in a nursing home at 5 AM. It is placed in a rack for pickup by the courier. The sample is picked up at 9 AM, delivered to the lab at 10:30 AM, and analyzed at 11:30 AM. Will the results be adversely affected by the handling of the specimen?
Yes.
Serum should be removed from cells within 1 hour of collection to prevent a DECREASE in glucose due to glycolysis.
When testing will be delayed, blood should be collected in a tube containing the antiglycolytic agent sodium fluoride, which preserves glucose for up to 3 days.
Other options are to centrifuge the blood and remove the serum or to use serum separator tubes.
What is glycated hemoglobin?
Hemoglobin A with glucose attached to the beta chains (Hemoglobin A1).
It is comprised of hemoglobin A1a, A1b, and A1c. Hemoglobin A1C is the largest fraction.
It is a useful indicator of long-term glucose control.
What specimen is required for an A1C?
EDTA whole blood
(Fasting is not required)
What are the most common methods for A1C in the U.S.?
Immunoassay or ion-exchange chromatography
What is the clinical significance of an elevated A1C?
It indicates poor glucose control over the past 2-3 months.
The A1C goal for diabetes is less than 7%.
Why is the A1C test invalid in a patient with hemoglobin S or C?
Because of the shortened life span of the RBCs.
A patient’s fasting plasma glucose is 99 mg/dL and her A1C is 12%. What do these results indicate?
Good short-term glucose control, but poor long-term control.
What is microalbuminuria?
Excretion of urinary albumin at a rate of 20-200 ug/minute or 30-300 mg/24 hr.
What is the significance of microalbuminuria?
It is highly predictive of diabetic nephropathy in type 1 and type 2 diabetes.
Early detection and tight glycemic control slow the progression of nephropathy.
How is microalbuminuria detected?
By immunochemical measurement of the albumin excretion rate on a 24-hr urine specimen using antibodies to human albumin.
What are the most common causes of hypoglycemia?
- inappropriate insulin production
- insulin injection
- ingestion of oral hypoglycemic agents
How is hypoglycemia diagnosed?
By the presence of Whipple’s triad:
1. plasma glucose <40 mg/dL
2. symptoms of hypoglycemia (nervousness, anxiety, neurologic abnormalities)
3. relief of symptoms by administration of glucose.
What happens when glucose levels drop too low?
CNS dysfunction
When are ketones present in the blood?
Whenever there is impaired carbohydrate metabolism with breakdown of fatty acids, such as uncontrolled diabetes mellitus, starvation, vomiting, or low carb diet.
The ketones are acetone, acetoacetic acid (acetoacetate or diacetic acid), and B-hydroxybutyric acid (B-hydroxybutyrate).
High levels lead to metabolic acidosis (ketoacidosis).
What reagent is most commonly used to detect ketones in blood and urine?
Sodium nitroprusside
Acetest tablets or reagent strip tests are used.
Sodium nitroprusside reacts with acetoacetic acid and to a lesser extent with acetone, but is insensitive to B-hydroxybutyric acid.
What is bilirubin?
the degradation product of heme.
Where is bilirubin produced?
in the reticuloendothelial cells following the breakdown of RBCS
Which protein transports bilirubin in the blood?
Albumin
Name the 3 types of bilirubin.
- Unconjugated (indirect) - bilirubin en route to the liver
- Conjugated (direct) - bilirubin once it goes through the liver - also known as bilirubin diglucuronide
- delta - covalently bonded to albumin and is only present with hepatic obstruction - it reacts as conjugated bilirubin in most methods.
Together these fractions make total bilirubin.
What happens to bilirubin in the liver?
It is conjugated with glucuronic acid by the enzyme uridyldiphosphate glucuronyl transferase (UDPG-T).
Following conjugation, it is excreted into the intestine via the bile duct and is reduced by bacteria to urobilinogen.
Urobilinogen is oxidized to urobilin, which gives stools their normal color.
What is the significance of clay-colored or light stools?
It is a sign of obstruction of the bile duct. Urobilin is not being produced because bilirubin is not reaching the intestines.
Which substances related to bilirubin metabolism is normally found in urine?
ONLY urobilinogen
Bilirubin should NOT be present in urine.
What urine chemistry abnormality is seen with complete obstruction of the biliary tract?
DECREASED urobilinogen
Which bilirubin fractions are analyzed in the laboratory?
Total and conjugated (direct)
The unconjugated (indirect) level is calculated by subtracting conjugated from total.
Compare the solubility of conjugated and unconjugated bilirubin.
Conjugated bilirubin is soluble in water; unconjugated bilirubin is not.
Both are soluble in alcohol.
Which form of bilirubin can be excreted in the urine?
ONLY conjugated bilirubin - it is not bound to protein, whereas unconjugated bilirubin is bound to albumin and is too large to be filtered through the glomeruli
What methods are used for determination of bilirubin levels?
Malloy- Evelyn and Jendrassik-Grof - both use a diazo reagent to react with bilirubin and produce colored azobilirubin.
Compare the Malloy-Evelyn and Jendrassik-Grof methods of bilirubin determination.
Malloy-Evelyn - carried out at an acid pH
Jendrassik-Grof - carried out at an alkaline pH
Conjugated bilirubin reacts without an accelerator (therefore called direct bilirubin); unconjugated bilirubin requires the addition of an accelerator (therefore called indirect bilirubin).
Name the accelerators for the Malloy-Evelyn and Jendrassik-Grof methods.
Malloy-Evelyn - methanol
Jendrassik-Grof - caffeine
Name a source of error that can DECREASE the level of bilirubin in a specimen.
Exposure to light.
Hemolysis - decreases by interfering with the reaction.
Name a source of error that can INCREASE the level of bilirubin in the specimen.
Lipemia
What is the normal range for total bilirubin in an adult?
0.2 - 1.0 mg/dL
What is the normal range for conjugated (direct) bilirubin in an adult?
<0.2 mg/dL
What is the normal range for unconjugated (indirect) bilirubin in an adult?
<0.8 mg/dL
How do normal values for bilirubin in a newborn compare to those in an adult?
Levels are higher in the newborn.
3-5 day old infant - 4-6 mg/dL
premature infant - 10-12 mg/dL
What would cause an increase in total bilirubin with a normal concentration of conjugated bilirubin?
PREhepatic jaundice
1. hemolytic transfusion reaction
2. hemolytic anemia
3. HDFN
What causes physiologic jaundice of the newborn?
Bilirubin metabolism is impaired because the newborn’s immature liver does not produce the enzyme required for bilirubin conjugation.
Bilirubin may remain elevated for 2 weeks.
In hemolytic disease of the fetus and newborn (HDFN), which bilirubin fraction is elevated and why?
Unconjugated, due to excessive breakdown of RBCs by maternal antibody.
What is the risk to the newborn from a high level of unconjugated bilirubin?
Unconjugated bilirubin (indirect) has a high affinity for brain tissue and causes necrosis (kernicterus). Without appropriate treatment, mental retardation, hearing deficits, or cerebral palsy may result.
At what level of bilirubin would an exchange transfusion be indicated in a neonate?
Once the unconjugated bilirubin reaches 20 mg/dL
What method may be used to determine neonatal bilirubin, but not adult bilirubin?
A non-invasive transcutaenous method in which a bilirubinometer performs multiple wavelength spectral analysis of the skin.
By taking readings at wavelengths that are absorbed by bilirubin, hemoglobin, and melanin, the method is able to determine total bilirubin without interference from hemoglobin or skin pigmentation.
This method cannot be used for patients over 1 month of age because of interfering dietary pigments, such as carotene.
Name 2 conditions in which conjugated bilirubin is elevated.
- hepatic jaundice
- posthepatic jaundice
What are the typical lab findings in posthepatic jaundice?
Increased total bilirubin
Increased conjugated bilirubin
Decreased urine urobilinogen
Clay-colored stools
Which disorder results in the highest levels of conjugated bilirubin?
Obstructive liver disease
How does hemolysis affect iron level?
Because of the high concentration of iron in hemoglobin, even minimal hemolysis will give falsely elevated results.
To minimize this effect, serum/plasma should be separated from RBCs within 1 hour of collection and even slightly hemolyzed specimens should not be analyzed.
How are iron levels affected by the time of day when the specimen is drawn?
Iron shows marked diurnal variation. Levels are approx. 30% higher in the morning.
Which protein transports iron?
Transferrin - it is normally 20-55% saturated with iron.
Where is most of the iron in the body?
In hemoglobin
Name 2 storage forms of iron.
- ferritin - primary storage form present in most cells and is a readily metabolized form of storage iron
- hemosiderin
How are the iron and total iron binding capacity (TIBC) affected in iron deficiency anemia?
Increased - serum iron and TIBC
What is TIBC?
an indirect measurement of transferrin
What is the most sensitive test for detection of iron deficiency anemia?
Serum ferritin - a decreased serum ferritin is almost always indicative of iron deficiency anemia
What is hereditary hemochromatosis?
the most common of the iron overload diseases - it causes the body to absorb and store too much iron
What tests are used to diagnose hereditary hemochromatosis?
Iron panel - serum iron, ferritin, TIBC, and transferrin saturation (percent saturation).
TS - determines how much iron is bound to transferrin, the protein that carries iron in the blood
After a 12 hour fast, the TIBC and SI are measured and the TS is calculated (SI/TIBC = TS)
The serum ferritin test shows the level of iron in the liver.
What test results are typical in hereditary hemochromatosis?
INCREASED: serum iron, ferritin, transferrin saturation
DECREASED: TIBC
Clinical significance of increased glucose levels (hyperglycemia)?
- diabetes mellitus
- other endocrine disorders
- acute stress
- pancreatitis
Clinical significance of decreased glucose levels (hypoglycemia)?
- insulinoma
- insulin-induced hypoglycemia
- hypopituitarism
What is the major source of cellular energy?
glucose
At room temperature, are glucose levels increased or decreased?
decreased
Which hormones decrease glucose levels?
Insulin
Action of insulin
responsible for entry of glucose into cells - increases glycogenesis
Which hormones increase glucose levels?
- glucagon
- cortisol
- epinenephrine
- growth hormone
- thyroxine
Action of glucagon on glucose
stimulates glycogenolysis & gluconeogenesis; inhibits glycolysis
Action of cortisol on glucose
insulin antagonist; increases gluconeogenesis
Action of epinephrine on glucose
promotes glycogenolysis and gluconeogenesis
Action of growth hormone on glucose
insulin antagonist
Action of thyroxine on glucose
increases glucose absorption from the GI tract; Stimulates glycogenolysis
Cause of gestational diabetes mellitus?
placental lactogen inhibits action of insulin
Typical laboratory findings in uncontrolled diabetes mellitus?
INCREASED:
1. blood glucose
2. urine glucose
3. urine SG
4. glycohemoglobin
5. ketones (blood and urine)
6. anion gap
7. BUN
8. Osmolality (serum and urine)
9. cholesterol
10. triglycerides
DECREASED:
1. bicarbonate
2. blood pH
What is metabolic syndrome?
a group of risk factors that seem to promote development of atherosclerotic cardiovascular disease and type 2 diabetes mellitus
What are the risk factors of metabolic syndrome?
INCREASED:
1. LDL-C
2. triglycerides
3. blood pressure
4. blood glucose
DECREASED:
1. HDL-C
Reference range for iron
M: 65-175 ug/dL
F: 50-170 ug/dL
Clinical significance of increased iron levels?
- iron overdose
- hemochromatosis
- sideroblastic anemia
- hemolytic anemia
- liver disease
Clinical significance of decreased iron levels?
iron deficiency anemia
What is necessary for hemoglobin synthesis and is transported by transferrin?
iron
What tube anticoagulants bind iron?
oxalate, citrate, and EDTA
Does hemolysis interfere with iron results?
yes
Why is an early morning specimen preferred for iron?
because of diurnal variation
Reference range for total iron binding capacity (TIBC)
250-425 ug/dL
Clinical significance of increased TIBC?
iron deficiency anemia
Clinical significance of decreased TIBC?
- iron overdose
- hemochromatosis
How is TIBC measured?
iron is added to transferrin. Excess removed. Iron content determined.
Reference range for % saturation or transferrin saturation
20-50%
Clinical significance of increased transferrin saturation?
- iron overdose
- hemochromatosis
- sideroblastic anemia
Clinical significance of decreased transferrin saturation?
iron deficiency anemia
Reference ranges for transferrin
200-360 mg/dL
Clinical significance of increased transferrin?
iron deficiency anemia
Clinical significance of decreased transferrin?
- iron overdose
- hemochromatosis
- chronic infections
- malignancies
How is transferrin saturation calculated?
100 x serum iron/TIBC
complex of apotransferrin and iron
transferrin
What is apotransferrin?
the protein that transports iron
Reference ranges for ferritin
M: 20-250 ug/L
F: 10-120 ug/L
Clinical significance of increased ferritin?
- iron overload
- hemochromatosis
- chronic infections
- malignancies
Clinical significance of decreased ferritin?
iron deficiency anemia
What stationary phase is used for the measurement of hemoglobin A1C by high performance liquid chromatography?
Cation exchanger
Impaired fasting glucose
plasma glucose >=100 but <126 mg/dL
Does Hgb F interfere with HPLC measurement of A1C?
Only if its concentration is greater than 30%
Does Hgb C interfere with HPLC measurement of A1C?
Not if it is completely separated on the chromatogram.
Does labile hemoglobin A1C interfere with HPLC measurement of A1C?
No, it produces a peak after Hgb F but before Hgb A1C.
Polarographic methods for glucose analysis are based upon what principle of measurement?
The rate of O2 depletion.
What enzyme is most specific for B-D-glucose?
Glucose oxidase
What coupling enzyme is used in the hexokinase method for glucose?
Glucose-6-phosphate dehydrogenase
What glucose method is subject to falsely low results caused by ascorbate?
Trinder glucose oxidase
Although glucose oxidase is specific for B-D-glucose, the coupling (indicator) reaction is prone to negative interference from ascorbate, uric acid, acetoacetic acid, and other reducing agents.
These compete with the chromogen for peroxide, resulting in less dye being oxidized to chromophore.
CSF glucose vs plasma glucose
The CSF glucose is usually 50-65% of the plasma glucose.
Clinical significance of low CSF glucose?
- bacterial or fungal meningitis
- malignancy in the CNS
- subarachnoid hemorrhage
- rheumatoid arthritis
- multiple sclerosis
Clinical significance of high CSF glucose?
A reflection of hyperglycemia and NOT CNS disease
In peroxidase-coupled glucose methods, what reagent complexes with the chromogen?
Phenol
POCTs for whole-blood glucose monitoring are based mainly on the use of what method?
amperometric detection
What effect does hematocrit have on POCT tests for whole-blood glucose monitoring?
Low hematocrit = increased readings
High hematocrit = lowers readings
Why does a high hematocrit lower the glucose?
because RBC glucose concentration is lower than plasma concentration
Other factors include: binding of oxygen to hemoglobin and the slower diffusion of glucose onto the solid phase - both of which occur when the hematocrit is high.
