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Clinical Chemistry 52: 903-904, 2006; 10.1373/clinchem.2005.065821
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(Clinical Chemistry. 2006;52:903-904.)
© 2006 American Association for Clinical Chemistry, Inc.

Letters to the Editor

Comparison of 3 Third-Generation Assays for Bio-intact Parathyroid Hormone

Nina Ljungdahl1, Mathias Haarhaus1,2, Cecilia Linder3 and Per Magnusson1,3,a

1 Bone and Mineral Metabolic Unit, Division of Clinical Chemistry, Department of Biomedicine and Surgery, Faculty of Health Sciences, Linköping, Sweden
2 Department of Nephrology, and, 3 Division of Clinical Chemistry, Department of Laboratory Medicine, Linköping University Hospital, Linköping, Sweden

aAddress correspondence to this author at: Division of Clinical Chemistry, Department of Biomedicine and Surgery, Faculty of Health Services, SE-58185 Linköping, Sweden. Fax 46-13-22-32-40; e-mail Per.Magnusson{at}lio.se.


To the Editor:

Current second-generation parathyroid hormone (PTH) assays suffer from interference by circulating PTH fragments. Newer assays that measure bio-intact PTH, i.e., the biologically active whole PTH molecule (1–84 PTH), have overcome this problem (1)(2). Our study was designed to evaluate and compare 3 commercial third-generation assays based on a chemiluminescence immunoassay, ELISA, and IRMA for bio-intact PTH and to compare these with a second-generation PTH assay.

EDTA-plasma samples were collected from 40 healthy blood donors (age range, 18–67 years; mean age, 39 years) and from 26 patients with predialysis chronic kidney disease (age range, 28–86 years; mean age, 65 years). Bio-intact PTH was measured by (a) a chemiluminescence immunoassay on the Nichols Advantage® platform (Nichols Institute Diagnostics) (2); (b) an IRMA using the Duo PTH reagents (Scantibodies Laboratory)(1); and (c) an ELISA using the Human BioActive Intact PTH reagents (Immutopics), which is a 2-site assay in which 2 goat polyclonal antibodies recognize epitopes within the midregion/carboxy terminus and at the initial amino terminus. For comparison, we performed a second-generation electrochemiluminescence PTH assay on the Elecsys 2010 platform (Roche Diagnostics). Ionized calcium and 25-hydroxyvitamin D (Nichols Advantage) were also measured in all samples.

Results for the healthy individuals are presented in Fig. 1 . The bio-intact PTH values measured by the IRMA and chemiluminescence immunoassay were, on average, 50% and 30% lower, respectively, than the PTH measured by the Elecsys. Regression analysis demonstrated significantly different slopes (P <0.001), but not intercepts, for both the IRMA and chemiluminescence immunoassay compared with the Elecsys assay. Most striking was the discordance between the ELISA and the IRMA, the chemiluminescence immunoassay, and the Elecsys assay. Comparison between the ELISA and Elecsys revealed a significantly different intercept (P <0.001). The results obtained with the third-generation IRMA and the chemiluminescence immunoassay also differed, despite the good correlation, and regression analysis demonstrated a significant difference for the slope (P <0.001) but not for the intercept. The finding of higher results with the chemiluminescence immunoassay concurs with the results reported by Boudou et al. (3), but immunoreactivity does not seem to be implicated in these differences(1)(2)(3). In addition to the apparent differences in assay design, other possible reasons for the lack of agreement between the assays investigated could be matrix effects and/or differences in standardization. The problems hampering immunoassay standardization are many and vary for different analytes, as has been reviewed by Stenman(4). Thus, the use of independent reference intervals for each of the bio-intact PTH assays is clinically important.


Figure 1
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  		Figure 1. Regression analysis for comparison of the 3 bio-intact PTH assays [IRMA (A), chemiluminescence immunoassay (CLIA; B), and ELISA (C)] vs the second-generation Elecsys assay, and for the comparison between the IRMA and the chemiluminescence immunoassay (D).

The data are for the healthy individuals (n = 40). The regression analysis equations are as follows: (A), y = 0.40x + 3.4 ng/L (r = 0.90; P <0.001); (B), y = 0.66x + 1.1 ng/L (r = 0.96; P <0.001); (C), y = –0.44x + 66 ng/L (r = –0.10; P, not significant); (D), y = 0.62x + 2.5 ng/L (r = 0.95; P <0.001).

We observed no significant differences between healthy individuals and patients with predialysis chronic kidney disease for 25-hydroxyvitamin D, but mean concentrations of bio-intact PTH (by chemiluminescence immunoassay; P <0.001) and ionized calcium (P <0.05) were higher in the patients. PTH measured by Elecsys was, on average, 75% higher than the measured bio-intact PTH in patients, consistent with the expected presence of high concentrations of circulating PTH fragments in patients with impaired renal function. In addition, the distribution of bio-intact PTH analyzed by the chemiluminescence immunoassay (mean, 71 ng/L; range, 5–227 ng/L) was wider in the patient group than in the healthy individuals [23 (11–50) ng/L].

Despite our strict adherence to the ELISA protocol and 3 independent runs with the same set of samples, this assay generated inconsistent results, which require additional methodologic studies to fully explain. We conclude that although the third-generation bio-intact PTH ELISA requires further evaluation, the chemiluminescence immunoassay and IRMA are comparable.


References

  1. Gao P, Scheibel S, D’Amour P, John MR, Rao SD, Schmidt-Gayk H, et al. Development of a novel immunoradiometric assay exclusively for biologically active whole parathyroid hormone 1–84: implications for improvement of accurate assessment of parathyroid function. J Bone Miner Res 2001;16:605-614.[CrossRef][ISI][Medline] [Order article via Infotrieve]
  2. Inaba M, Nakatsuka K, Imanishi Y, Watanabe M, Mamiya Y, Ishimura E, et al. Technical and clinical characterization of the bio-PTH (1–84) immunochemiluminometric assay and comparison with a second-generation assay for parathyroid hormone. Clin Chem 2004;50:385-390.[Abstract/Free Full Text]
  3. Boudou P, Ibrahim F, Cormier C, Chabas A, Sarfati E, Souberbielle J-C. Third- or second-generation parathyroid hormone assays: a remaining debate in the diagnosis of primary hyperparathyroidism. J Clin Endocrinol Metab 2005;90:6370-6372.[Abstract/Free Full Text]
  4. Stenman U-H. Immunoassay standardization: is it possible, who is responsible, who is capable?. Clin Chem 2001;47:815-820.[Free Full Text]




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