Super resolution imaging of protein-protein interactions

Abstract

This disclosure provides methods and compositions for detecting intramolecular and intermolecular interactions, such as protein-protein interactions. These methods detect such interactions at sub-diffraction distances, and thus are referred to as super-resolution detection and imaging methods.

1 . A composition comprising:
 (a) a first conjugate comprising a first binding partner linked to a first oligonucleotide that comprises a half-docking domain and a stability domain;   (b) a second conjugate comprising a second binding partner linked to a second oligonucleotide that comprises a half-docking domain and a stability domain;   wherein the stability domains of the first conjugate is complementary to the stability of the second conjugate, and wherein the first half-docking domain and the second half-docking domain form a full docking domain when the stability domain of the first conjugate binds to the stability domain of the second conjugate; and   (c) a detectably-labeled imager strand that is complementary to the full docking domain.   
     
     
         2 . The composition of  claim 1 , wherein the first binding partner is an antibody or antigen-binding antibody fragment that binds to a first target protein, and the second binding partner is an antibody or antigen-binding antibody fragment that binds to a second target protein. 
     
     
         3 . The composition of  claim 1 , wherein the first binding partner is an antibody or antigen-binding antibody fragment that binds to a first epitope on a target protein, and the second binding partner is an antibody or antigen-binding antibody fragment that binds to a second epitope on the target protein. 
     
     
         4 . The composition of  claim 1 , wherein the first half-docking domain has a length of 5-15 nucleotides and the second half-docking domain has a length of 5-15 nucleotides. 
     
     
         5 .- 6 . (canceled) 
     
     
         7 . The composition of  claim 1 , wherein the detectably-labeled imager strand has a length of 10-30 nucleotides. 
     
     
         8 .- 12 . (canceled) 
     
     
         13 . The composition of  claim 1 , wherein the detectably-labeled imager strand comprises a fluorescent label. 
     
     
         14 . The composition of  claim 1 , wherein the first binding partner is linked to the first oligonucleotide via a streptavidin-biotin binding pair, and the second binding partner is linked to the second oligonucleotide via a streptavidin-biotin binding pair. 
     
     
         15 . A complex comprising the composition of  claim 1 , a first target protein bound to the first binding partner, and a second target protein bound to the second binding partner. 
     
     
         16 . (canceled) 
     
     
         17 . A plurality of compositions, wherein each of the compositions comprises the composition of  claim 1 , and wherein at least one of the compositions of the plurality comprises a detectably-labeled imager strand that is spectrally distinct from other imager strands in the plurality spectrally-distinct labels. 
     
     
         18 . (canceled) 
     
     
         19 . A plurality of compositions, wherein each of the compositions comprises the composition of  claim 1 , and wherein at least one of the compositions of the plurality has a blinking frequency that is distinct from other compositions in the plurality. 
     
     
         20 . A method of detecting a complex of two protein targets in a sample, the method comprising:
 contacting a sample with the composition of  claim 1 , wherein the first binding partner binds to one of the two targets, and the second binding partner binds to the other of the two targets; and   detecting presence or absence of the complex in the sample.   
     
     
         21 .- 26 . (canceled) 
     
     
         27 . A method of detecting an intramolecular interaction on a target protein in a sample, the method comprising:
 contacting a sample that comprises a target molecule with the composition of  claim 1 , wherein the first binding partner binds to a first epitope of the target protein, and the second binding partner binds to a second epitope of the target protein; and   detecting presence or absence of the intramolecular interaction in the sample.   
     
     
         28 .- 30 . (canceled) 
     
     
         31 . A composition comprising
 (a) a first conjugate comprising a first binding partner linked to a first oligonucleotide that comprises a half-docking domain and a stability domain;   (b) a second conjugate comprising a second binding partner linked to a second oligonucleotide that comprises a half-docking domain and a stability domain;   wherein the stability domains of the first conjugate is complementary to the stability of the second conjugate, and wherein the first half-docking domain and the second half-docking domain form a first full docking domain when the stability domain of the first conjugate binds to the stability domain of the second conjugate;   (c) a third conjugate comprising a third binding partner linked to a third oligonucleotide that comprises a half-docking domain and a stability domain,   wherein the stability domains of the first conjugate is complementary to the stability domain of the third conjugate, and wherein the first half-docking domain and the third half-docking domain form a second full docking domain when the stability domain of the first conjugate binds to the stability domain of the third conjugate;   (d) a first detectably-labeled imager strand that is complementary to the first full docking domain; and   (e) a second detectably-labeled imager strand that is complementary to the second full docking domain.   
     
     
         32 . The composition of  claim 1 , wherein the detectably-labeled imager strand comprises a 5′ domain that is complementary to the first half-docking domain, a 3′ domain that is complementary to the second half-docking domain, and a linker domain located between the 5′ domain and the 3′-domain. 
     
     
         33 . The composition of  claim 1 , wherein the first oligonucleotide comprises a spacer domain between the first half-docking domain and the stability domain of the first conjugate, and the second oligonucleotide comprises a spacer domain between the second half-docking domain and the stability domain of the second conjugate. 
     
     
         34 . The composition of  claim 32 , wherein the 5′ domain of the detectably-labeled imager strand has a length of 5-10 nucleotides, the 3′ domain of the detectably-labeled imager strand has a length of 5-10 nucleotides, and/or the linker domain of the detectably-labeled imager strand has a length of 1-5 nucleotides. 
     
     
         35 . The composition of  claim 32 , wherein the linker domain comprises thymine (T) nucleotides. 
     
     
         36 . The composition of  claim 32 , wherein the linker domain comprises a TT sequence. 
     
     
         37 . A complex comprising the composition of  claim 1  and a target protein comprising a first epitope and a second epitope, wherein the first binding partner is bound to the first epitope, and the second binding partner is bound to the second epitope.