Analytical and clinical validation of a time-resolved immunofluorometric assay (TR-IFMA) for canine C-reactive protein in serum.
A time-resolved immunofluorometric assay (TR-IFMA) was developed for the determination of C-reactive protein (CRP) in canine serum. CRP was isolated from canine acute-phase serum by affinity chromatography on agarose coupled with phosphorylethanolamine. This isolated dog CRP was used as standard to calibrate the assay. Intra-assay and inter-assay coefficients of variation were in the ranges 5.3-7.1% and 4.8-13.3%, respectively. Accuracy, evaluated by adding 2 and 10 microg/ml of CRP to serum samples, provided recoveries of 99.9% and 106.8%. High correlation was found between CRP measurements by TR-IFMA and a by commercial enzyme-linked immunosorbent assay (R2 = 0.98). The limit of detection for the TR-IFMA method was 0.000067 microg/ml and the measurement of CRP in serial dilutions of acute-phase dog sera generated curves with the same slope as the one constructed with purified CRP. The TR-IFMA provides a precise, accurate and highly sensitive assay for CRP determination in dog samples. CRP levels in dogs with different diseases ranged between 10.2 and 210.7 microg/ml and were significantly higher than those observed in healthy dogs (< 7.1 microg/ml).
Parra
M D
MD
Department of Animal Medicine and Surgery, Faculty of Veterinary Medicine, University of Murcia, 30100 Murcia, Spain. mdparra@um.es
Tuomola
M
M
Cabezas-Herrera
J
J
Cerón
J J
JJ
D000210:Acute-Phase Reaction / Q000097:blood; D000818:Animals; D002097:C-Reactive Protein / Q000032:analysis*; D004283:Dog Diseases / Q000097:blood*; D004285:Dogs; D004797:Enzyme-Linked Immunosorbent Assay / Q000662:veterinary; D015200:Fluoroimmunoassay / Q000379:methods / Q000662:veterinary*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D002097:C-Reactive Protein