Mouse Anti-PLAU Recombinant Antibody (clone mAb-112) (CAT#: PABL-306)

Figure 1 Functional effects of antibodies on in vitro plasminogen activation initiated by pro-uPA or active uPA.

0.25 nM pro-uPA or 0.25 nM uPA was incubated without (light blue dots) antibody or with 5 nM (pink dots), 10 nM (dark blue dots), 20 nM (yellow dots), 40 nM (green dots), 80 nM (red dots) and 160 nM (black dots) of mAb-112, mAb-PUK, mAb-12E6B10, mAb-101 or anti-PAI-1 mAb-2, as indicated. After incubation for 30 min at room temperature, plasminogen was added to 0.5 µM and S-2251 to 0.5 mM. Substrate hydrolysis was followed at 37°C by measuring the absorbance at 405 nm. Pro-uPA or uPA was omitted in controls (grey curves).

Botkjaer, K. A., Fogh, S., Bekes, E. C., Chen, Z., Blouse, G. E., Jensen, J. M.,... & Declerck, P. J. (2011). Targeting the autolysis loop of urokinase-type plasminogen activator with conformation-specific monoclonal antibodies. Biochemical Journal, 438(1), 39-51.

Figure 2 Effect of the antibodies on cell surface-associated plasminogen activation.

U937 cells were washed with glycine-HCl, pH 3, 0.1 M NaCl to remove pro-uPA and uPA bound to the cell surface. One nM pro-uPA or uPA alone or pre-incubated with monoclonal antibodies were added to the cells together with 0.4 µM α2-antiplasmin and 0.2 µM plasminogen. A, Example of a time course experiment with pro-uPA preincubated alone (black) or with mAb-PUK at 0.2 nM (blue), 2 nM (yellow), 20 nM (green) or 200 nM (red). B, IC50 plots of inhibition of cell surface-associated plasminogen activation by mAb-101 (black), mAb-112 (green) and mAb-PUK (red) in an assay with pro-uPA. C, IC50 plots of inhibition by mAb-101 (black), mAb-112 (green) and mAb-12E106B (yellow) in an assay with active uPA. All curves in B and C were fit to a hyperbolic decay equation, which provided approximate IC50-values.

Botkjaer, K. A., Fogh, S., Bekes, E. C., Chen, Z., Blouse, G. E., Jensen, J. M.,... & Declerck, P. J. (2011). Targeting the autolysis loop of urokinase-type plasminogen activator with conformation-specific monoclonal antibodies. Biochemical Journal, 438(1), 39-51.

Figure 3 Inhibitory effects of antibodies on tumour cell invasion in vitro and dissemination in vivo.

A, Matrigel invasion assay, in which PC-Hi/diss cells were allowed to invade the matrix barrier towards chemoattractants present in chicken embryonic fibroblast conditioned medium in the presence of 333 nM control IgG, mAb-112, mAb-PUK or mAb-12E6B10 or 0.1 TIU/ml aprotinin. The data are presented as percentage of control and are means ± SEM. ** and ***, p<0.005 and 0.0001, respectively. B, Tumour cell escape assay, in which PC-hi/diss cells were allowed to escape the initial collagen droplet and invade a fibrin-enriched collagen matrix. Antibodies and aprotinin were incorporated in final concentrations of 167 nM and 0.1 TIU/ml, respectively. The invasion index was calculated as the number of escaped tumour cells multiplied by the distance invaded. The data are presented as percentage of control and are means ± SEM. * and **, p<0.05 and 0.01, respectively. C and D, Spontaneous dissemination assay in chick embryos. PC-hi/diss cells were inoculated on the chorioallantoic membrane (CAM) of chicken embryos and allowed to form primary tumours, which were excised and weighed after 7 days of growth (C). The numbers of disseminated PC-hi/diss cells were determined by Alu qPCR in the portions of the CAM distal to the site of primary tumour formation (D). Data are presented as numbers of human cells per 10⁶ chicken cells and are means ± SEM.