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Quantification of Nucleosomes in Serum,,,by the Cell Death Detection ELISAplus

ting native anti-histone antibodies to the normal immunoreagent containing biotinylated anti-histone antibodies. With increasing concentrations of the native anti-histone antibodies, the absorbance values declined in a dosedependent manner indicating that only complexes of DNA and histones bound to peroxidase-labeled anti- DNA antibodies and biotinylated anti-histone antibodies were detected by the CDDE. DNA, histones, nucleosomes, anti-histone antibodies or anti-DNA antibodies alone do not alter the measured values (16) (Figure 2).



Imprecision

The imprecision, as a criterion for the reliability of the ELISA, ranged in the intra-assay comparison (n=10) between 3.0% and 4.1 %, in the inter-assay comparison (n=14) between 8.6% and 13.5 % [16].

Stabilization

In the serum and plasma of ten individuals, which were stored under varying conditions (storage time, temperature, addition of a stabilizer), the best stability was obtained for serum which has been treated with 10mM EDTA (pH 8) and stored at -20 C or -80 C [16].

Long-term stability

Serum samples which were treated with EDTA and stored at -20 C showed a good long-term stability at low and high absorbance levels, even after six months. This was indicated by a coefficient of variation between 4.2% and 9.2 % [16] (Figure 3).



Preanalytic handling of the samples

The standardization of the preanalytic handling of the samples was essential for the reliable quantification of nucleosomes in serum. Whereas hemolysis and prolongation of the period between ven
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