C-reactive protein (CRP) is an extremely powerful prognostic marker for a wide variety of diseases. In human medicine immunoturbidimetric assays (ITAs) are replacing the ELISA test due to their high degree of accuracy, high-throughput and low turnaround time. Until now, a canine-specific ELISA test has represented the gold standard in veterinary medicine for measuring serum CRP. Even though there are contrasting data concerning the cross-reactivity of human and canine CRP, a previous study has nevertheless demonstrated that a commercially available human ITA is reliable for measuring canine CRP without method modification. Since many automated analyzers are furnished with proprietary standardized reagents, the aim of this study was to verify whether a commercially available human ITA (Olympus system reagent, CRP ITA, Olympus, Italia) different from the previous validated assay is also reliable for accurately measuring canine CRP. The CRP concentration of 35 canine sera collected from either healthy dogs or dogs with infectious and neoplastic diseases was assayed by ELISA and ITA on an automated chemistry analyzer (Olympus AU 400). The ELISA test was carried out following the manufacturer’s instructions while the ITA measurement was initially performed using the settings and calibration of the human assay. The results, which were compared using Passing and Bablok regression analysis, showed that the ITA underestimated canine CRP both with an absolute and a proportional inaccuracy. To overcome this pitfall, the ITA was repeated after calibration of the automated analyzer with a standard specimen obtained by pooling three canine sera with high CRP concentrations. The standard was assayed 6 times in the same run using the ELISA test. The CV was 5.8% and the mean concentration of the 6 replicates was assumed to be the actual concentration of the standard. The modified assay (ITA-c) was used to evaluate the same 35 samples in duplicate. The results were again compared with the ELISA test by Passing and Bablok regression analysis which showed a good correlation (y = 1.18x – 0.69). Moreover, two samples with 23.7mg/dl and 21.7 mg/dl of CRP, respectively, were serially diluted and assayed in duplicate. The ITA-c was linear in the verified range (y = 0.9988x + 0.2 and y = 0.9982x + 0.07). In conclusion, this study confirms the evidence that any commercially available human CRP ITA should be validated before its routine application in dogs. Moreover, the partial cross-reactivity between human and canine CRP allowed us to reliably measure canine CRP after calibration of the ITA with a standard specimen having a known concentration of the analyte.
F. Gentilini, D. Mancini, F. Dondi, L. Ingrà, M.E. Turba, M. Forni, et al. (2005). Validation of a human immunoturbidimetric assay for measuring canine C-reactive protein. GLASGOW : s.n.
Validation of a human immunoturbidimetric assay for measuring canine C-reactive protein
GENTILINI, FABIO;MANCINI, DANILO;DONDI, FRANCESCO;INGRA', LAURA;TURBA, MARIA ELENA;FORNI, MONICA;FAMIGLI BERGAMINI, PAOLO
2005
Abstract
C-reactive protein (CRP) is an extremely powerful prognostic marker for a wide variety of diseases. In human medicine immunoturbidimetric assays (ITAs) are replacing the ELISA test due to their high degree of accuracy, high-throughput and low turnaround time. Until now, a canine-specific ELISA test has represented the gold standard in veterinary medicine for measuring serum CRP. Even though there are contrasting data concerning the cross-reactivity of human and canine CRP, a previous study has nevertheless demonstrated that a commercially available human ITA is reliable for measuring canine CRP without method modification. Since many automated analyzers are furnished with proprietary standardized reagents, the aim of this study was to verify whether a commercially available human ITA (Olympus system reagent, CRP ITA, Olympus, Italia) different from the previous validated assay is also reliable for accurately measuring canine CRP. The CRP concentration of 35 canine sera collected from either healthy dogs or dogs with infectious and neoplastic diseases was assayed by ELISA and ITA on an automated chemistry analyzer (Olympus AU 400). The ELISA test was carried out following the manufacturer’s instructions while the ITA measurement was initially performed using the settings and calibration of the human assay. The results, which were compared using Passing and Bablok regression analysis, showed that the ITA underestimated canine CRP both with an absolute and a proportional inaccuracy. To overcome this pitfall, the ITA was repeated after calibration of the automated analyzer with a standard specimen obtained by pooling three canine sera with high CRP concentrations. The standard was assayed 6 times in the same run using the ELISA test. The CV was 5.8% and the mean concentration of the 6 replicates was assumed to be the actual concentration of the standard. The modified assay (ITA-c) was used to evaluate the same 35 samples in duplicate. The results were again compared with the ELISA test by Passing and Bablok regression analysis which showed a good correlation (y = 1.18x – 0.69). Moreover, two samples with 23.7mg/dl and 21.7 mg/dl of CRP, respectively, were serially diluted and assayed in duplicate. The ITA-c was linear in the verified range (y = 0.9988x + 0.2 and y = 0.9982x + 0.07). In conclusion, this study confirms the evidence that any commercially available human CRP ITA should be validated before its routine application in dogs. Moreover, the partial cross-reactivity between human and canine CRP allowed us to reliably measure canine CRP after calibration of the ITA with a standard specimen having a known concentration of the analyte.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
