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Establishment of murine monoclonal antibodies against murine urokinase receptor raised in urokinase receptor knock-out mice

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  • Ebbe Ranne
  • Gunilla H. Hansen
  • Behrendt, Niels
  • Hélène Solberg
  • Jørgen K. Larsen
  • Keld Danø

The urokinase plasminogen activator receptor (uPAR) plays a central role during cancer invasion and metastasis by localizing and focalizing the uPA mediated proteolytic activity to the cell surface.We have previously raised monoclonal antibodies to the human uPAR (Renne, E. et al (1991) FEBS Lett. 288, 233-236) and used them for different analysis of human uPAR. None of the antibodies showed any cross-reactivity to murine uPAR. In order to study the murine uPAR further we immunized uPAR knock-out mice ( Bugge, T.H. et al (1995) J. Biol. Chem. 270, 16886-16894) with human soluble uPAR, based on the assumption that it might be possible to raise antibodies to epitopes highly conserved between human and mouse. From one fusion we obtained 22 stable hybridomas. Two of the hybridomas produced antibodies that cross-react with the murin uPAR as analysed by immunoblotting using Tx-114 detergent phase extract from a murine cell line. Further analysis of the antibodies showed that the react with an epitop on domain 2+3 of uPAR and only under non-reducing conditions of the antigen. The antibodies also showed useful for immunoaffinity purification of soluble murin uPAR as well as staining of uPAR on cells as demonstrated by flow cytometry analysis. In conclusion , we have demonstrated that it is possible to raise antibodies to highly conserved epitopes between human and mouse uPAR by immunizing uPAR knock-out mice with human uPAR. The antibodies generated might be a useful tool for demonstrating the occurence and amount of rnurine uPAR in different biological samples but further studies are needed.

OriginalsprogEngelsk
TidsskriftFibrinolysis and Proteolysis
Vol/bind11
Udgave nummerSUPPL. 3
Antal sider1
ISSN1369-0191
StatusUdgivet - 1 dec. 1997

ID: 241481146