Heme derivatives: test procedures

6 ASCP MLS practice questions on Heme derivatives: test procedures, part of Chemistry. The first 6 are below; the first 5 with answers and explanations (tap Show answer after you have chosen), the rest in Learn with a free account.

1. Question 1 · Easy

   In diazo methods, bilirubin couples with diazotized sulfanilic acid to form:

   1. ABilirubin glucuronide
   2. BBiliverdin
   3. CAzobilirubin
   4. DUrobilinogen

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   Answer: C · Azobilirubin

   The diazo reagent splits bilirubin at its central carbon bridge, and each half couples with the diazonium salt to give azobilirubin. The color formed is proportional to the bilirubin present and is read photometrically. Conjugated bilirubin reacts quickly in water, while unconjugated bilirubin needs an accelerator. This reaction is the basis of the Jendrassik-Grof and Malloy-Evelyn methods.

   • A: Bilirubin glucuronide is made by the liver during conjugation, not by the test.
   • B: Biliverdin forms when bilirubin is oxidized, as in bilirubin oxidase methods.
   • D: Urobilinogen is made by gut bacteria and reacts with Ehrlich's reagent.

2. Question 2 · Hard

   A technologist runs a Jendrassik-Grof total bilirubin on a jaundiced newborn whose bilirubin is almost all unconjugated, but leaves out the caffeine-sodium benzoate. The total bilirubin result will be:

   1. AUnchanged, because alkaline tartrate frees bilirubin
   2. BFalsely high, because free diazo reagent absorbs
   3. CFalsely low, near the direct bilirubin value
   4. DUnchanged, because unconjugated bilirubin reacts directly

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   Answer: C · Falsely low, near the direct bilirubin value

   Without an accelerator, only conjugated and delta bilirubin react within the timed reading, which is the definition of direct bilirubin. Albumin-bound unconjugated bilirubin reacts very slowly, so most of it is missed. In a newborn with mainly unconjugated bilirubin, the total would be far too low and treatment could be missed. A total close to the direct value in a visibly jaundiced infant should prompt a check of the reagents.

   • A: Alkaline tartrate only changes the color of azobilirubin already formed.
   • B: Ascorbic acid destroys the excess diazo reagent, so it adds no color.
   • D: Unconjugated bilirubin reacts slowly without an accelerator, which is why it is called indirect.

3. Question 3 · Hard

   A newborn's bilirubin sample was sent in a clear tube and left on a bright windowsill for 2 hours before testing. The result is 9.2 mg/dL (157 µmol/L); a transcutaneous reading at the time of the draw was 14.8 mg/dL (253 µmol/L). The best action is:

   1. ARecollect, protecting the new tube from light
   2. BRepeat the assay on the same tube
   3. CReport a direct bilirubin instead
   4. DReport the result with a light-exposure note

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   Answer: A · Recollect, protecting the new tube from light

   Light breaks down bilirubin in the tube, much as phototherapy does in the skin, so a sample left in bright light gives a falsely low result. The large gap from the transcutaneous value fits this error. Repeating the test on the same tube cannot recover the lost bilirubin, and a falsely low value could delay phototherapy or exchange transfusion. A new sample should be collected in an amber tube or one wrapped in foil, and tested promptly.

   • B: The bilirubin already destroyed in the tube cannot be recovered.
   • C: Direct bilirubin is also light-sensitive and does not replace the total.
   • D: A result known to be falsely low should not be reported, even with a comment.

4. Question 4 · Medium

   In the Jendrassik-Grof total bilirubin method, caffeine-sodium benzoate is added to:

   1. AForm the diazonium salt from sulfanilic acid
   2. BRelease unconjugated bilirubin from serum albumin
   3. CDestroy excess diazo reagent at the end
   4. DTurn azobilirubin blue for reading at 600 nm

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   Answer: B · Release unconjugated bilirubin from serum albumin

   Unconjugated bilirubin is tightly bound to albumin and poorly soluble in water, so it reacts slowly with the diazo reagent. Caffeine-sodium benzoate frees it so that it reacts as fast as conjugated bilirubin, and the result is total bilirubin. Direct bilirubin is measured in a separate tube without the accelerator. Ascorbic acid then stops the reaction, and alkaline tartrate shifts the color to blue for reading near 600 nm.

   • A: Sulfanilic acid and sodium nitrite together form the diazo reagent.
   • C: Ascorbic acid stops the reaction by destroying excess diazo reagent.
   • D: Alkaline tartrate, added at the end, turns azobilirubin blue.

5. Question 5 · Medium

   Direct spectrophotometric bilirubin, read near 454 nm with a second wavelength to correct for hemoglobin, is used for newborns but not adults. Why?

   1. AConjugated bilirubin in adults does not absorb at 454 nm
   2. BAlbumin-bound bilirubin in adults absorbs no light
   3. CAdult hemoglobin is not corrected by the second reading
   4. DAdult serum has carotenoids that absorb near 454 nm

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   Answer: D · Adult serum has carotenoids that absorb near 454 nm

   Bilirubin absorbs strongly near 454 nm, and a second reading, usually at 540 nm, subtracts the contribution of hemoglobin. Newborn serum has almost no carotenoids or other lipochromes, so the 454 nm reading reflects bilirubin. Adult serum carries dietary carotenoids that also absorb near 454 nm and would falsely raise the result. This is why direct bilirubinometers are used only for neonatal samples.

   • A: Conjugated and unconjugated bilirubin both absorb near 454 nm.
   • B: Bilirubin bound to albumin still absorbs light near 454 nm.
   • C: The second reading corrects for hemoglobin of any type, adult or fetal.

6. Question 6 · Easy

   Why should a neonatal bilirubin sample be protected from light?

   1. ALight hemolyzes the red cells in the sample
   2. BLight changes the sample's pH and protein binding
   3. CLight raises bilirubin, giving falsely high results
   4. DLight breaks down bilirubin, giving falsely low results

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