Artifactual prolongation of activated partial thromboplastin time with PEGylated compounds in silica-based assays

Conjugation of polyethylene glycol (PEG) to another molecule prolongs its half-life in the body, but has a potential to artifactually increase the activated partial thromboplastin time (aPTT) as measured with certain assays. Studies conducted in-house at Bristol-Myers Squibb using the STA-PTT Automate 5 assay, the routine assay used to measure aPTT, demonstrated prolongation of aPTT in plasma samples spiked in vitro with 40-kDa branched PEG (PEG40) conjugated compounds or PEG40 alone, but not in samples spiked with vehicle or non-PEGylated compound, suggesting that the interference is because of the PEG40 moiety. To investigate the cause of this phenomenon, cynomolgus monkey and rat plasma samples were spiked with different concentrations of PEG40 and the aPTT was measured using different proprietary assays. With one exception, prolongation of aPTT was observed with all assays containing silica as the contact activator. No changes in aPTT were noted in assays using kaolin as a contact activator. The findings indicated that the observed prolongation of aPTT is largely because of interference of PEG40 with the silica, but other features of the reagent mixture may also influence aPTT times.