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Letters |
1st Lab. Clin. Chem., Clin. Chem. and Enzymol. Section, Spedali Civili, 25125 Brescia, Italy
a Author for correspondence.
To the Editor:
We evaluated the Opus Troponin I assay (Behring Diagnostics, Westwood, MA), a two-site sandwich, fluorogenic ELISA that uses two goat polyclonal antibodies directed against different protein segments unique to cardiac troponin I (cTnI) (1). Pipetting, incubations, measurements, and data-reduction steps are performed on the Opus analyzer; the first test result requires 20 min. The assay measures concentrations of cTnI in serum as great as ~135 µg/L.
The calibration appeared to be stable for at least 4 weeks. Serial dilution of a human serum sample with a high concentration of cTnI showed no significant curvature when the curve obtained was tested for linearity [quadratic regression: y = -0.25 + 96.1x + 6.31x2, with the coefficient of x2 not significantly different from 0 (P = 0.27)] (2). Linear regression analysis of these data confirmed the high linearity of the response (r = 0.9998). The minimum detectable cTnI concentration, assessed by 10 replicate measurements of a human serum containing no detectable cTnI concentration and defined as the cTnI value corresponding to the fluorescence signal 3 SD greater than the mean found for this serum, was estimated as 0.38 µg/L. The Opus analyzer, however, reports results <0.50 µg/L as "<0.5 µg/L."
Assay reproducibility was tested by assaying in duplicate, once a day for 10 days, two serum samples with concentrations distributed over the measuring range and the three kit controls containing human cTnI (3). Analysis of variance showed within-run CVs between 3.4% and 7.2% and total CVs between 5.6% and 13.0%. No interferences were detected in assays of lipemic (triglycerides <10 g/L) or hemolyzed (hemoglobin <2.5 g/L) specimens; concentrations of bilirubin >50 mg/L spuriously increased the reported cTnI concentrations in serum.
To compare the Opus assay with the cTnI Pasteur immunoenzymometric assay (Sanofi Diagnostics Pasteur, Marnes-la-Coquette, France; performed manually according to the manufacturer's current protocol (4)), we assayed 85 unselected individual serum samples with detectable cTnI concentration (>0.04 µg/L as measured with the Pasteur assay). The correlation was good (r = 0.959), but the data showed considerable scatter (Sy|x = 10.6 µg/L), the Opus results being relatively higher within the mid-range of values but equal to the Pasteur results at concentrations outside the limit of linearity of the instrument. As a whole, the Opus assay led to approximately 10-fold higher values than the Pasteur assay (linear regression equation: Opus = 11.1 Pasteur + 1.5 µg/L). Differences in the calibration materials and in the ways their theoretical values are assigned in the two methods may partially explain this observation. When the three calibrators supplied with the Pasteur kit (S1, S2, and S3 with assigned cTnI concentrations of 0.13, 0.55, and 1.56 µg/L, respectively) were determined with the Opus assay, the results were as follows: 1.21, 4.39, and 11.3 µg/L, respectively.
We used the Opus procedure to measure the concentration of cTnI in sera
from subjects in four groups: (a) 46 apparently healthy
people, ages 26–82 years; (b) 21 patients with a typical
history of myocardial infarction (MI) of <8 h duration, not treated
with thrombolytic therapy; (c) 8 patients with severe
skeletal muscle damage [total creatine kinase (CK) values 10 240 to
226 000 U/L] but no apparent cardiac injury; and (d) 39
consecutive dialysis patients with no evidence of myocardial injury.
CK-MB mass concentrations were determined with the Magic Lite assay
(Ciba Corning Diagnostic Corp., Medfield, MA). No cTnI was detected in
the healthy subjects (all values <0.5 µg/L). In patients with MI,
cTnI peaked at 20.8 ± 8.1 h (range, 8–33 h) after the onset
of chest pain, reaching a mean peak concentration of 165 µg/L (range,
3.3–1674 µg/L). Comparing their upper reference limits, we noted a
higher relative increase at peak in cTnI than in CK-MB mass
concentration (mean, 330- vs 47-fold). Furthermore, unlike the short
duration of increased CK-MB concentrations, cTnI remained diagnostic
over all the period tested, i.e., from the arrival at the hospital
until 96 h after onset of pain, being still >0.5 µg/L in 16 of
18 samples obtained at day 4 post-MI (Table 1
).
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Consistent with the manufacturer's declared cross-reactivity of <0.007% with skeletal troponin I (1), cTnI was undetectable in all but one of the patients with severe skeletal muscle damage or chronic renal failure. That patient had a slightly increased cTnI (1.0 µg/L) while receiving continuous ambulatory peritoneal dialysis; clinical evaluation, however, revealed no evidence of myocardial injury.
We conclude that the Opus method has the potential to become a valuable aid in specific detection of myocardial injury.
Acknowledgments
We thank Istituto Behring, Milan, Italy, for supplying reagents for the cTnI assay.
References
The following articles in journals at HighWire Press have cited this article:
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  R. Labugger, J. A. Simpson, M. Quick, H. A. Brown, C. E. Collier, I. Neverova, and J. E. Van Eyk Strategy for Analysis of Cardiac Troponins in Biological Samples with a Combination of Affinity Chromatography and Mass Spectrometry Clin. Chem., June 1, 2003; 49(6): 873 - 879. [Abstract] [Full Text] [PDF]
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D. J. Newman, Y. Olabiran, W. D. Bedzyk, S. Chance, E. G. Gorman, and C. P. Price Impact of Antibody Specificity and Calibration Material on the Measure of Agreement between Methods for Cardiac Troponin I Clin. Chem., June 1, 1999; 45(6): 822 - 828. [Abstract] [Full Text] [PDF]
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M. Panteghini, R. Bonora, F. Pagani, F. Buffoli, and C. Cuccia Rapid, Highly Sensitive Immunoassay for Determination of Cardiac Troponin I in Patients with Myocardial Cell Damage Clin. Chem., August 1, 1997; 43(8): 1464 - 1465. [Full Text]
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