US 12,276,654 B2
Chemical probe-dependent evaluation of protein activity and uses thereof
Raymond E. Moellering, Chicago, IL (US); and Gang Li, Chicago, IL (US)
Assigned to The University of Chicago, Chicago, IL (US)
Appl. No. 15/733,113
Filed by The University of Chicago, Chicago, IL (US)
PCT Filed Nov. 21, 2018, PCT No. PCT/US2018/062231
§ 371(c)(1), (2) Date May 19, 2020,
PCT Pub. No. WO2019/104155, PCT Pub. Date May 31, 2019.
Claims priority of provisional application 62/589,946, filed on Nov. 22, 2017.
Prior Publication US 2020/0355673 A1, Nov. 12, 2020
Int. Cl. G01N 33/50 (2006.01); A61K 31/7088 (2006.01); C07K 14/00 (2006.01); C07K 16/44 (2006.01); C12P 19/34 (2006.01); G01N 33/542 (2006.01)
CPC G01N 33/5035 (2013.01) [A61K 31/7088 (2013.01); C07K 14/003 (2013.01); C07K 16/44 (2013.01); C12P 19/34 (2013.01); G01N 33/542 (2013.01); C07K 2317/622 (2013.01); C07K 2318/20 (2013.01); G01N 2458/10 (2013.01)] 20 Claims
OG exemplary drawing
 
1. A method for evaluating two or more target proteins of interest from the same family in a specified functional form, the method comprising:
(i) contacting a sample comprising or suspected of comprising the two or more proteins of interest with a molecular construct comprising: a targeting group operatively linked to a retrieval tag; wherein the targeting group comprises a small molecule probe that specifically binds and covalently links to the specialized functional form of the two or more target proteins of interest;
(ii) contacting the sample with at least two antibody-oligo constructs, wherein at least one of the constructs comprises a first antibody operatively linked to a first oligo and at least a second construct comprises a second antibody operatively linked to a second oligo; wherein the first antibody specifically binds to one of the two or more target proteins of interest and the second antibody specifically binds to the other of the two or more target proteins of interest;
(iii) contacting the sample with a second molecular construct comprising a retrieval tag binder operatively linked to a retrieval oligo;
(iv) incubating the sample under conditions sufficient for the ligation or annealing of the first oligo to the retrieval oligo when the first and retrieval oligos are in close proximity to each other and ligation of the second oligo to the retrieval oligo when the second and retrieval oligos are in close proximity to each other; and
(v) detecting the ligated or annealed first and retrieval oligo and the ligated or annealed second and retrieval oligo.