Mechanism of interference by haemolysis in the cardiac troponin T immunoassay

Ann Clin Biochem 2006;43:49-56
doi:10.1258/000456306775141687
© 2006 Association for Clinical Biochemistry

 

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Original Articles


Ravinder Sodi,
Simon M Darn,
Andrew S Davison,
Anthony Stott and
Alan Shenkin


Department of Clinical Biochemistry and Metabolic Medicine, Royal Liverpool and Broadgreen University Hospital, Prescot Street, Liverpool, L7 8XP, UK;
Department of Clinical Biochemistry and Metabolic Medicine, Royal Liverpool and Broadgreen University Hospital, Prescot Street, Liverpool, L7 8XP, UK;
Department of Clinical Biochemistry and Metabolic Medicine, Royal Liverpool and Broadgreen University Hospital, Prescot Street, Liverpool, L7 8XP, UK;
Department of Clinical Biochemistry and Metabolic Medicine, Royal Liverpool and Broadgreen University Hospital, Prescot Street, Liverpool, L7 8XP, UK;
Department of Clinical Biochemistry and Metabolic Medicine, Royal Liverpool and Broadgreen University Hospital, Prescot Street, Liverpool, L7 8XP, UK


Background: The cardiac troponins have been shown to be sensitiveand specific biochemical markers of myocardial infarction andhighly prognostic for future adverse events in patients withacute coronary syndromes. There have been reports suggestingthat haemolysis causes a negative interference in the cardiactroponin T (cTnT) assay but the mechanism(s) involved remainunknown. Here we show the effects of haemolysis and haemoglobinper se on the cTnT assay.

Methods: The effect of haemolysis was studied by the additionof prepared haemolysate to serum samples with known and clinicallyrelevant cTnT levels. The effect of haemoglobin was studiedby the addition of haemoglobin of increasing concentrationsand noting its effect on the level of cTnT measured. The effectof putative proteases was determined indirectly by incubatingsamples with spiked cTnT with various protease inhibitors andobserving the changes in the measured cTnT levels.

Results: The results show that both haemolysis, which is therelease of haemoglobin and corpuscular contents, and haemoglobinitself negatively interfere in the cTnT assay in a concentration-dependentmanner, although the former had a greater magnitude of effect.On haemolysis, indirect evidence suggests that proteases arereleased which degrade the cTnT in serum, thus causing the decreasedlevels detected. Pepstatin A, a reversible inhibitor of asparticproteinases, effectively inhibited the loss of cTnT in serumat 37°C and pH 7.4 over a 48-h period. We found that ata haemoglobin level of 0.75 g/L, cTnT declined by more than10% of the initial concentration, suggesting that falsely decreasedlevels due to haemolysis may significantly affect the clinicalutility of the assay.

Conclusions: Haemolysis, haemoglobin per se and possibly proteolysisplay a role in the negative interference in cTnT assays. Measuresto reduce this interference must be implemented.


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