Degradation of Cardiac Troponin I in Serum Complicates Comparisons of Cardiac Troponin I Assays

Abstract

Background: Up to a 20-fold variation in serum cardiac troponin I (cTnI) concentration may be observed for a given patient sample with different analytical methods. Because more limited variation is seen for control materials and for purified cTnI, we explored the possibility that cTnI was present in altered forms in serum. Methods: We used four recombinantly engineered cTnI fragments to study the regions of cTnI recognized by the Stratus®, Opus®, and ACCESS® immunoassays. The stability of these regions in serum was analyzed with Western blot. Results: The measurement of several control materials and different forms of purified cTnI using selected commercial assays demonstrated five- to ninefold variation. Both the Stratus and Opus assays recognized the N-terminal portion (NTP) of cTnI, whereas the ACCESS assay recognized the C-terminal portion (CTP) of cTnI. Incubation of recombinant cTnI in normal human serum produced a marked decrease in cTnI concentration as determined with the ACCESS, but not the Stratus, immunoassay. Western blot analysis of the same samples using cTnI NTP- and CTP-specific antibodies demonstrated preferential degradation of the CTP of cTnI. Conclusions: The availability of serum cTnI epitopes is markedly affected by the extent of ligand degradation. The N-terminal half of the cTnI molecule was found to be the most stable region in human serum. Differential degradation of cTnI is a key factor in assay-to-assay variation.

Evaluation of a high‐sensitivity assay for measurement of canine and feline serum cardiac troponin I

“…The healthy control animals in this study had cTnI concentrations comparable to those of healthy animals reported in other studies . Although results of different assays cannot be directly compared, many studies using different cTnI assays report similar values for healthy animals, whereas the assays differ considerably (up to 20‐fold) in the higher concentration ranges . As release of higher concentrations of cTnI invariably also leads to a higher proportion of circulating modified cTnI, it has been speculated that some assays perform better at detecting modified forms than others.…”

Comparison of Canine Cardiac Troponin I Concentrations as Determined by 3 Analyzers

“…[17][18][19] The use of purified complexed cTnI (to cardiac troponin T or C) might have been useful in this study; however, similar differences in analytical methods between purified cTnI and patient cTnI values have been noted in human medicine. 20 Although the lack of a common calibrator among analyzers may be a major reason for the large differences in results seen both in this study and in human studies, other factors may be at play. Additional explanations for the differences among analyzers as well as the differences among purified samples and clinical samples include different target amino acids for each analyzer, the variable complexed or noncomplexed forms of cTnI recognized by the antibodies in each assay, and modification (degradation or phosphorylation) of circulating cTnI after myocardial injury that variably influences the detection of cTnI by different analyzers.…”

“…Additional explanations for the differences among analyzers as well as the differences among purified samples and clinical samples include different target amino acids for each analyzer, the variable complexed or noncomplexed forms of cTnI recognized by the antibodies in each assay, and modification (degradation or phosphorylation) of circulating cTnI after myocardial injury that variably influences the detection of cTnI by different analyzers. 1,19,20 This latter point may explain why the fold differences among analyzers were greater at higher cTnI concentrations; perhaps the Access is better able to detect modified forms of cTnI after cardiac injury than the Biosite and Stratus. Such cTnI modifications have been shown in rat models and in human beings; however, this hypothesis has not been investigated in dogs.…”

“…[17][18][19] Several studies in human medicine have compared analyzers, and although results obtained on different analyzers correlate well, a large (up to 20-fold) difference in the absolute values obtained between some machines has been observed. [13][14][15]17,20,21 The use of and the potential applications for cTnI analysis are growing in veterinary medicine; however, the inability to compare values obtained on different analyzers has limited collaborative studies between institutions to this point. Reference ranges for dogs have been developed for several different analyzers using the 95th percentile as the upper end of the range, and the established ranges are similar for 3 of these analyzers (Biosite Triage Meter a ,0.05-0.12 ng/mL, Dade-Behring Stratus b ,0.03-0.07 ng/mL, and Beckman-Coulter Access AccuTnI c 0.01-0.11 ng/mL).…”

Preliminary Investigation of Cardiac Troponin I Concentration in Cows with Common Production Diseases

“…45,46 Variations in the measured cTnI concentration occur when the same samples are analyzed with different immunoassays. 46 The absolute measured cTnI concentration is not comparable among different assays. Therefore, the results of this study are only valid for measurement of bovine serum cTnI concentrations with the i-STAT immunoassay and cannot be compared to other immunoassays.…”