Method for the detection of a binding partner of a multispecific binder

Abstract

Herein is reported a method for the detection of free antigen of a multispecific antibody in a sample, whereby the antigen to be detected can be specifically bound by a first binding site of the multispecific antibody, comprising the step of incubating a sample comprising free antigen and multispecific antibody with an anti-idiotypic antibody, which specifically binds to a second binding specificity of the bispecific antibody, which is different from the first binding specificity, whereby the anti-idiotypic antibody is bound to a solid phase.

1 - 10 . (canceled) 
     
     
         11 . A method for the determination of the presence or amount of free antigen of a multispecific antibody in a sample comprising the free antigen and the multispecific antibody, wherein the antigen can be specifically bound by a first binding specificity of the multi specific antibody, comprising the steps of:
 (A) incubating the sample comprising the free antigen and the multispecific antibody with a second antigen that can be specifically bound by a second binding specificity of the multispecific antibody, which is different from the first binding specificity, to form a second antigen-multispecific antibody complex and   (B) removing the second antigen-multi specific antibody complex from the sample to produce a multispecific antibody-depleted sample prior to the determination of the presence or amount of the antigen.   
     
     
         12 . The method according to  claim 11 , wherein the second antigen is conjugated to a solid phase. 
     
     
         13 . The method according to  claim 11 , wherein the second antigen is conjugated to a paramagnetic bead. 
     
     
         14 . The method according to  claim 11 , further comprising
 (C) determining the presence or amount of free antigen in the multispecific-antibody depleted sample.   
     
     
         15 . The method according to  claim 14 , wherein determining the presence or amount of the free antigen in the multispecific antibody depleted sample comprises the following steps:
 (i) incubating the multispecific antibody-depleted sample with a capture antibody that specifically binds to the antigen to form a capture antibody-antigen complex, and   (ii) correlating the formed capture antibody-antigen complex to the amount of the free antigen in the sample.   
     
     
         16 . The method according to  claim 14 , wherein determining of the presence or amount of free antigen in the multispecific antibody depleted sample comprises the following steps:
 (i) incubating the multi specific antibody-depleted sample with a capture antibody that specifically binds to the antigen to form a capture antibody-antigen complex,   (ii) incubating the capture antibody-antigen complex with a tracer antibody to form a capture antibody-antigen-tracer antibody complex, wherein the capture antibody and the tracer antibody bind to non-overlapping epitopes on the antigen, and   (iii) correlating the formed capture antibody-antigen-tracer antibody complex to the amount of the antigen in the sample.   
     
     
         17 . The method according to  claim 14 , wherein determining the presence or amount of the free antigen in the multispecific antibody depleted sample comprises the following steps:
 (i) incubating the multispecific antibody-depleted sample with a capture antibody that specifically binds to the antigen to form a capture antibody-antigen complex,   (ii) incubating the capture antibody-antigen complex with a tracer antibody to form a capture antibody-antigen-tracer antibody complex, wherein the capture antibody and the tracer antibody bind to non-overlapping epitopes on the antigen,   (iii) incubating the capture antibody-antigen-tracer antibody complex with a detection antibody comprising a detectable label, wherein the detection antibody specifically binds to the tracer antibody at an epitope outside the variable domains of the tracer antibody, and   (iv) correlating the formed capture antibody-antigen-tracer antibody complex to the amount of the free antigen in the sample.   
     
     
         18 . The method according to  claim 11 , wherein the multispecific antibody is a bispecific antibody that has a first binding specificity that specifically binds to a first antigen or first epitope on an antigen and that has a second binding specificity that specifically binds to a second antigen or to a second epitope on the antigen. 
     
     
         19 - 21 . (canceled)