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Clin Chem. 2010 Sep;56(9):1424-31. doi: 10.1373/clinchem.2009.141663. Epub 2010 Jul 12.

Dry-reagent double-monoclonal assay for cystatin C.

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Department of Biotechnology, University of Turku, Tykistökatu 6A, 6th Floor, FIN-20520 Turku, Finland.



Cystatin C is a low molecular weight cysteine proteinase inhibitor whose plasma or serum concentrations have been shown to be better correlated with glomerular filtration rate than serum creatinine concentrations. Routine assays for cystatin C are based on use of polyclonal antibodies and immunoturbidimetric and nephelometric designs. This study aimed to develop a double-monoclonal immunoassay for cystatin C.


We tested functionality of 42 2-site antibody combinations involving 7 monoclonal antibodies with recombinant and plasma cystatin C. We developed a heterogeneous assay using 2 antibodies selected to give the best analytical performance. The assay used a dilution step and was based on a dry-reagent, all-in-one immunoassay concept with time-resolved fluorometry. The assay was performed on an automated immunoanalyzer in single wells that contained all the required assay components. We used heparin-derived plasma samples for methodological evaluation of the assay.


From a relative epitope map involving 7 cystatin C-specific antibodies, we selected a pair of antibodies for a 2-site sandwich-type dry-reagent assay. Total assay time was 15 min, and 10 microL of a 100-fold diluted sample was used. The analytical detection limit (background + 3SD) and functional detection limit (CV 20%) were 0.01 mg/L and 0.02 mg/L, respectively. Within-run and total assay imprecision were <4.7% and <5.6% (at 0.84-3.2 mg/L), respectively, and plasma recoveries of added cystatin C were 94%-110%. Regression analysis with the Roche particle-enhanced immunoturbidimetric method yielded the following (SD): slope, 1.391 (0.029); y-intercept, -0.152 (0.045) mg/L; S(y logical or, bar belowx)=0.294 mg/L (n=131).


The developed assay enables rapid and reliable measurement of cystatin C.

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