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IMMUNO-BASED RETARGETED ENDOPEPTIDASE ACTIVITY ASSAYS

  • US 20190324041A1
  • Filed: 07/08/2019
  • Published: 10/24/2019
  • Est. Priority Date: 03/13/2009
  • Status: Active Grant
First Claim
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1. A method of detecting retargeted endopeptidase activity, the method comprising the steps of:

  • a) treating a cell from an established cell line with a sample comprising a retargeted endopeptidase, wherein the cell from an established cell line is susceptible to retargeted endopeptidase activity by a retargeted endopeptidase;

    b) isolating from the treated cell line a SNAP-25 component comprising a SNAP-25 cleavage product having a carboxyl-terminus at the P1 residue of the BoNT/A cleavage site scissile bond;

    c) contacting the SNAP-25 component with an α

    -SNAP-25 antibody, wherein the α

    -SNAP-25 antibody binds an epitope comprising a carboxyl-terminus at the P1 residue of the BoNT/A cleavage site scissile bond from a SNAP-25 cleavage product; and

    d) detecting the presence of an antibody-antigen complex comprising the α

    -SNAP-25 antibody and the SNAP-25 cleavage product;

    wherein detection by the antibody-antigen complex is indicative of retargeted endopeptidase activity, wherein the α

    -SNAP-25 antibody binds an epitope comprising a carboxyl-terminus at the P1 residue from the BoNT/A cleavage site scissile bond from a SNAP-25 cleavage product and wherein the isolated α

    -SNAP-25 antibody has a VH CDR1 of SEQ ID NO;

    93, a VH CDR2 of SEQ ID NO;

    96, a VH CDR3 of SEQ ID NO;

    100, a VL CDR1 of SEQ ID NO;

    105, a VL CDR2 of SEQ ID NO;

    110, and a VL CDR3 of SEQ ID NO;

    115.

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