42 and 3.23 ng/mL, respectively (Table 3). The assay sensitivity was estimated as the assay cut point multiplied by the minimum sample dilution factor. The assay sensitivity was therefore estimated to be 28.3 and 64.5 ng/mL for anti-velaglucerase alfa and anti-imiglucerase

antibodies, respectively. This approach to determine the cut point is therefore supported by the assay sensitivities estimated for this assay, which are higher (28.3 ng/mL and 64.5 ng/mL) than the assay sensitivities of the antibody screening assay (33.4 ng/mL and 65.6 ng/mL). It is recognized that assay signal responses can vary over time and between assay runs (Mire-Sluis et al., 2004), particularly for assays utilizing radiolabeled reagents, and therefore the cut point CPM value for this assay may be adjusted to compensate for radiolabel decay. In MG132 order to normalize inter-assay variability, the cut point CPM can be adjusted, when necessary, relative to the calibration curve slope and y-intercept. The least squares line fit to the high-purity, monoclonal antibody-based calibration curve data, using well-characterized known concentrations of antibody, provides a reliable and consistent method for calculating the uncertainty in assay determinations. This procedure normalizes the cut point for inter-assay changes in CPM that may

occur from reagent radiolabel decay, radioautolysis and/or assay handling variability as well as allowing for changes in non-specific binding and for changing www.selleckchem.com/products/Avasimibe(CI-1011).html assay readouts over time. For all the confirmatory assays (IgA, IgM and IgE), Sitaxentan precision and linearity were determined according to guidelines (FDA, 2001, ICH, 2005 and EMEA, 2009) and are described in Table 4 and Table 5. The positive cut points for both the anti-velaglucerase alfa and anti-imiglucerase IgA and IgM confirmatory assays (Table 4 and Table 5) were determined from the mean blank (buffer only) result from 59 and 68 assays, respectively, for both velaglucerase alfa and imiglucerase. The positive cut points were calculated by the signal-to-noise approach

as 10 times the mean blank. Of note, these calculated cut point values were below or near the instrument limit of detection. A ratio of 2.0, indicating a 2-fold increase in signal over baseline, has been described in the literature as a clinically acceptable criterion for an anti-drug antibody-positive sample (Miller et al., 2001). Patient sera are therefore defined as positive for anti-velaglucerase alfa or imiglucerase IgA or IgM antibodies if the signal of the timepoint is greater than or equal to the respective cut point and if the ratio of the timepoint signal to the pre-infusion baseline signal is greater than or equal to 2.0. For the IgE assays, the assay cut point was established as the mean plus 1.645 standard deviation of assay values obtained from treatment-naïve patient serum samples as recommended by Mire-Sluis et al. (2004).