Human Anti-GP6 Recombinant Antibody (clone hGP 5C4) (CAT#: HPAB-0133CQ)

Figure 2 The ability of hGP 5C4 Fab to inhibit GPVI binding to collagen was monitored in an ELISA-based assay.

Adhesion of the Fc-GPVI-nt-consisting of the extracellular domain of GPVI and the Fc part of an IgG-to immobilised collagen was investigated in the presence of different anti-GPVI antibody Fab fragments (20 µg/ml). Binding of the Fc part without the GPVI receptor domain served as control. The binding is visualised with a secondary antibody directed to the Fc part of Fc-GPVI-nt labelled with peroxidase. Peroxidase (PE) is finally detected by an ELISA system measuring the absorption photometrically at 450 nm. hGP 5C4 prevents Fc-GPVI-nt binding to collagen, whereas 4C9 could not prevent collagen interaction with GPVI. The means ± SEM are shown.

Figure 3 The binding of different antibodies to stable GPVI-expressing CHO cells was measured by FACS.

The binding of the specific antibodies to control CHO cells served as control. After incubation with the primary antibodies CHO cells were incubated with anti-IgG rat antibodies labelled with PE. Specific PE fluorescence was determined in a Becton Dickenson FACScalibur device.

Figure 4 The inhibition of collagen-mediated aggregation and ATP release by hGP 5C4 was tested in human platelets.

In simultaneous experiments, ATP release was measured form these platelets. Collagen induced marked release of ATP from intracellular stores. hGP 5C4 potently inhibited this collagen-induced ATP release. The ATP release is given in % of control release. The means ± SEM are shown.

Figure 5 The influence of hGP 5C4 on bleeding time was tested in human blood ex vivo.

Human blood was incubated with hGP 5C4 Fab in 10 to 20 fold therapeutic concentrations (5 µg/ml) and bleeding time was determined with a PFA-100 device. Bleeding time was compared to ReoProR in equivalent concentrations. There was no significant prolongation of the bleeding time with hGP 5C4 Fab compared to control blood, whereas Reopro led to a maximal prolongation of bleeding time.