Case Study: A Dark-Brown Serum Sample

This case study was originally published in Clinical Chemistry

A 65-year-old woman with congestive heart failure presented for a mitral valve replacement. She experienced excessive bleeding and thrombosis during surgery, with subsequent hemolytic anemia.

Serum samples were abnormally dark. The hemolysis and icteric indices were not increased above the laboratory's threshold for electrolyte measurement; however, the lipemia index was. Upon visual inspection, the sample was not milky or turbid in appearance.



  1. What is the most likely cause of the dark color observed in the patient sample?

  2. What factors can influence the measurement of lipemic index?

  3. What laboratory tests might identify the cause of the dark color? How should samples with abnormal color be handled to verify that the color is not interfering with analytical methods?


Methemoglobin, myoglobin, and methemalbumin can cause abnormal dark color in patient samples. “Lipemia” was measured at 660- and 700-nm wavelengths; therefore any compounds that absorb light in this range will result in an increased lipemic index. In this patient, methemoglobin was always less than 7.5 percent (cooximetry), myoglobin was not detected (gel filtration chromatography and mass spectrometry), but an interference causing falsely increased results by immunoturbidimetry was observed. Haptoglobin was undetectable (nephelometry). Methemalbumin, formed when albumin binds excess hemin (oxidized free heme), was markedly increased (multiwavelength spectrophotometry), consistent with intravascular hemolysis. Chemistry analyses were performed on both undiluted serum and serial dilutions to confirm acceptable analyte recovery and rule out potential interference.

Authors: Maria Willrich, Ph.D., Craig Wittwer, Nikola Baumann, Ph.D., and Darci Block, Ph.D.

Kelley Luedke (@kschrib)

Kelley Luedke

Kelley Luedke is a Marketing Channel Manager at Mayo Clinic Laboratories. She is the principle editor and writer of Insights and leads social media and direct marketing strategy. Kelley has worked at Mayo Clinic since 2013. Outside of work, you can find Kelley running, traveling, playing with her kitty, and exploring new foods.