Multiplex immuno screening assay

A method for detecting at least two target antibodies in a biological sample comprising:

• (a) contacting a first solid support bound to an AGT substrate covalently coupled to a first fusion protein comprising an AGT polypeptide having a O6-alkylguanine-DNA alkyltransferase activity and a first epitope that is recognized by a first target antibody with the biological sample;
• (b) contacting a second solid support bound to an AGT substrate covalently coupled to a second fusion protein comprising an AGT polypeptide having a O6-alkylguanine-DNA alkyltransferase activity and a second epitope that is recognized by a second target antibody, but not by said first target antibody with the biological sample;
• and
• (c) detecting the presence or absence of the two target antibodies.

A method for covalently coupling an AGT polypeptide having a O6-alkylguanine-DNA alkyltransferase activity, on a functionalized solid support, comprising the following steps:

• a) activating the said functionalized solid support,
• b) adding a substrate of said AGT polypeptide, said substrate being suspended in a buffer containing between 0 and 20% of DMSO, in appropriate conditions so that the substrate is covalently attached to said support,
• c) contacting said AGT polypeptide with the substrate-coated support of step b) in a PBS/DTT buffer,
• wherein unbound molecules are washed out after steps b) and c).