A 54-year-old man with obstructive jaundice from a tumour of the pancreatic head has blood drawn into a lithium heparin tube, which is spun and separated within 20 minutes. The plasma is visibly icteric, with total bilirubin 410 µmol/L (24 mg/dL) (reference below 21 µmol/L), and is not hemolysed or lipemic. On the main analyzer, glucose by glucose oxidase with a peroxidase-coupled (Trinder) color reaction is 4.7 mmol/L (85 mg/dL). The backup analyzer, using hexokinase with glucose-6-phosphate dehydrogenase (G6PD), gives 6.0 mmol/L (108 mg/dL) on the same plasma a few minutes later. Both analyzers passed QC today. Which result should be reported, and why?

Hexokinase result; bilirubin uses up peroxide in the Trinder step

Glucose oxidase result; it is the reference glucose method

Glucose oxidase result; hexokinase also phosphorylates fructose

Neither result; glycolysis has lowered both, so recollect

Which enzymatic method is generally taught as the reference method for plasma glucose because of its high specificity?

Glucose oxidase with an oxygen electrode

Sodium fluoride does not fully stop the fall in glucose during the first 1 to 2 hours after collection. Why?

It blocks enolase, a late step, and takes time to act

It is neutralized by the oxalate in the same tube

It works only after the sample has been refrigerated

It inhibits glycolysis in white cells but not red cells

Glucose methods, specimens and interferences

13 DOH Abu Dhabi (HAAD) lab licensing practice questions on Glucose methods, specimens and interferences, part of Clinical Chemistry. The first 13 are below; the first 5 with answers and explanations (tap Show answer after you have chosen), the rest in Learn with a free account.

Question 1 · Hard
A 54-year-old man with obstructive jaundice from a tumour of the pancreatic head has blood drawn into a lithium heparin tube, which is spun and separated within 20 minutes. The plasma is visibly icteric, with total bilirubin 410 µmol/L (24 mg/dL) (reference below 21 µmol/L), and is not hemolysed or lipemic. On the main analyzer, glucose by glucose oxidase with a peroxidase-coupled (Trinder) color reaction is 4.7 mmol/L (85 mg/dL). The backup analyzer, using hexokinase with glucose-6-phosphate dehydrogenase (G6PD), gives 6.0 mmol/L (108 mg/dL) on the same plasma a few minutes later. Both analyzers passed QC today. Which result should be reported, and why?
1. AGlucose oxidase result; it is the reference glucose method
2. BGlucose oxidase result; hexokinase also phosphorylates fructose
3. CHexokinase result; bilirubin uses up peroxide in the Trinder step
4. DNeither result; glycolysis has lowered both, so recollect
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Answer: C · Hexokinase result; bilirubin uses up peroxide in the Trinder step
Both results come from the same plasma, separated within 20 minutes, so glycolysis cannot explain the gap; it would lower both results equally. In the Trinder step, peroxidase uses the hydrogen peroxide formed from glucose to oxidize a dye, and reducing substances such as bilirubin, ascorbic acid and uric acid compete for that peroxide, so the glucose oxidase result reads falsely low. Hexokinase with G6PD is the reference method: hexokinase can also phosphorylate fructose and mannose, but G6PD oxidizes only glucose-6-phosphate, so the method stays specific. The hexokinase result is reported, and icteric samples above the limit in the glucose oxidase insert should be sent to the hexokinase method.
- A: Hexokinase, not glucose oxidase, is the reference method for glucose.
- B: The G6PD step reacts only with glucose-6-phosphate, so fructose does not raise the hexokinase result.
- D: Glycolysis would lower both results equally, and this plasma was separated within 20 minutes.
Question 2 · Easy
Which enzymatic method is generally taught as the reference method for plasma glucose because of its high specificity?
1. AGlucose dehydrogenase with PQQ
2. BHexokinase
3. CGlucose oxidase-peroxidase
4. DGlucose oxidase with an oxygen electrode
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Answer: B · Hexokinase
Hexokinase phosphorylates glucose with ATP, and glucose-6-phosphate dehydrogenase (G6PD) then oxidizes the glucose-6-phosphate while reducing NADP+ to NADPH, read at 340 nm. The second step acts only on glucose-6-phosphate, so few substances interfere. For this reason hexokinase is the basis of the classical reference method. Most large chemistry analyzers use it for routine plasma glucose.
- A: GDH with PQQ also reacts with maltose and xylose, so it is not specific.
- C: The peroxidase step is prone to interference from reducing agents such as ascorbic acid.
- D: The oxygen-electrode method is specific but is not the accepted reference method.
Question 3 · Easy
In the laboratory glucose dehydrogenase method that uses NAD+, the product measured at 340 nm is:
1. ANADPH
2. BNADH
3. CHydrogen peroxide
4. DQuinoneimine dye
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Answer: B · NADH
Glucose dehydrogenase oxidizes beta-D-glucose to gluconolactone and reduces NAD+ to NADH. The rise in absorbance at 340 nm is proportional to the glucose concentration, and mutarotase may be added to speed the reaction. The NAD-linked enzyme is very specific for glucose. Point-of-care GDH strips that use the PQQ cofactor are different: they also react with maltose and xylose.
- A: NADPH is formed in the hexokinase-G6PD method, not with NAD-linked GDH.
- C: Hydrogen peroxide is a product of glucose oxidase.
- D: A quinoneimine dye is the color formed in the Trinder peroxidase step.
Question 4 · Medium
Sodium fluoride does not fully stop the fall in glucose during the first 1 to 2 hours after collection. Why?
1. AIt blocks enolase, a late step, and takes time to act
2. BIt is neutralized by the oxalate in the same tube
3. CIt works only after the sample has been refrigerated
4. DIt inhibits glycolysis in white cells but not red cells
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Answer: A · It blocks enolase, a late step, and takes time to act
Fluoride inhibits enolase near the end of glycolysis, and it takes time to enter the cells. Until then, the earlier enzymes such as hexokinase and phosphofructokinase keep using glucose, so the level keeps falling for the first hour or two. Tubes with an acidic citrate buffer inhibit these early steps at once. For diagnostic glucose, use such a tube or put the sample in an ice-water slurry and separate it within 30 minutes.
- B: Oxalate is the anticoagulant and does not neutralize fluoride.
- C: Fluoride works at room temperature; cooling only slows glycolysis further.
- D: Fluoride inhibits glycolysis in red cells as well as white cells.
Question 5 · Medium
A ward sends only a grey-top tube (sodium fluoride/potassium oxalate) for glucose, lactate, ethanol and potassium. Which result cannot be reported?
1. AEthanol
2. BLactate
3. CPotassium
4. DGlucose
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Answer: C · Potassium
The potassium oxalate anticoagulant adds a large amount of potassium, so the result would be falsely high. Fluoride also inhibits many enzymes, so fluoride plasma is unsuitable for enzyme activities and for some enzymatic methods, such as urease-based urea. Glucose, lactate and ethanol are routinely measured in fluoride tubes, because fluoride stops cells from consuming glucose and making lactate and prevents microbes from forming alcohol. A separate lithium heparin or serum tube is needed for potassium.
- A: Fluoride tubes are used for blood alcohol because they prevent microbial fermentation.
- B: Fluoride stops red cells from producing lactate, so it is the preferred tube.
- D: Fluoride-oxalate plasma is the standard sample for glucose.
Question 6 · Hard
A laboratory changes from sodium fluoride/potassium oxalate tubes to citrate-buffered fluoride tubes with a dry additive. In paired samples, all analyzed 2 hours after collection, fasting glucose now reads about 0.3 mmol/L (5 mg/dL) higher. What is the best explanation?
1. AFluoride in the old tube inhibited the hexokinase reagent
2. BThe buffer lyses red cells, which release extra glucose
3. CIts acid citrate buffer stops glycolysis almost at once
4. DCitrate gives a positive interference in the hexokinase step
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Question 7 · Easy
Once serum is separated from the cells, glucose in an unhemolyzed, sterile sample is stable at room temperature for about:
1. A7 days
2. B8 hours
3. C30 minutes
4. D2 hours
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Question 8 · Easy
In the glucose oxidase reaction, glucose and oxygen form gluconic acid and:
1. AHydrogen peroxide
2. BNADPH
3. CCarbon dioxide
4. DGlucose-6-phosphate
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Question 9 · Easy
A clinic sends its glucose samples to a laboratory 3 hours away. Which tube best limits glycolysis during transport?
1. ALight blue top: 3.2% sodium citrate
2. BGrey top: sodium fluoride/potassium oxalate
3. CLavender top: dipotassium EDTA
4. DGold top: clot activator and gel
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Question 10 · Medium
In the hexokinase glucose method, specificity for glucose comes mainly from:
1. APhosphorylation of glucose by hexokinase
2. BRemoval of protein before the reaction
3. COxidation of glucose-6-phosphate by G6PD
4. DReading the absorbance at 340 nm
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Question 11 · Medium
In the oxygen-electrode glucose oxidase method, molybdate with iodide (or ethanol with catalase) is added to:
1. ARemove ascorbic acid that would reduce the dye
2. BStop the peroxide formed from giving back oxygen
3. CSupply the oxygen that the enzyme consumes
4. DConvert alpha-glucose quickly to beta-glucose
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Question 12 · Medium
Two hours after a 75 g glucose load, compared with venous plasma glucose drawn at the same time, capillary plasma glucose is usually:
1. AHigher, as tissues extract glucose from the blood
2. BLower, because capillary red cells use more glucose
3. CLower, because tissue fluid dilutes capillary blood
4. DThe same, because both are measured as plasma
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Question 13 · Hard
An outpatient's fasting blood was drawn into a gel serum tube at 07:30 and spun at 10:00 after transport in a warm van. Glucose is 6.7 mmol/L (121 mg/dL); the diabetes cut-off is 7.0 mmol/L (126 mg/dL). The best action is:
1. ACorrect it for 2.5 hours of loss and report the new value
2. BRecollect, with the tube on ice and spun within 30 minutes
3. CReport it as impaired fasting glucose with no comment
4. DRerun the same serum on a second analyzer to confirm it
Answer and explanation in LearnFree account

Glucose methods, specimens and interferences

Which enzymatic method is generally taught as the reference method for plasma glucose because of its high specificity?

In the laboratory glucose dehydrogenase method that uses NAD+, the product measured at 340 nm is:

Sodium fluoride does not fully stop the fall in glucose during the first 1 to 2 hours after collection. Why?

A ward sends only a grey-top tube (sodium fluoride/potassium oxalate) for glucose, lactate, ethanol and potassium. Which result cannot be reported?

A laboratory changes from sodium fluoride/potassium oxalate tubes to citrate-buffered fluoride tubes with a dry additive. In paired samples, all analyzed 2 hours after collection, fasting glucose now reads about 0.3 mmol/L (5 mg/dL) higher. What is the best explanation?

Once serum is separated from the cells, glucose in an unhemolyzed, sterile sample is stable at room temperature for about:

In the glucose oxidase reaction, glucose and oxygen form gluconic acid and:

A clinic sends its glucose samples to a laboratory 3 hours away. Which tube best limits glycolysis during transport?

In the hexokinase glucose method, specificity for glucose comes mainly from:

In the oxygen-electrode glucose oxidase method, molybdate with iodide (or ethanol with catalase) is added to:

Two hours after a 75 g glucose load, compared with venous plasma glucose drawn at the same time, capillary plasma glucose is usually:

An outpatient's fasting blood was drawn into a gel serum tube at 07:30 and spun at 10:00 after transport in a warm van. Glucose is 6.7 mmol/L (121 mg/dL); the diabetes cut-off is 7.0 mmol/L (126 mg/dL). The best action is: