US 9,809,848 B2
Simultaneous detection of target protein and target nucleic acids in a single cell
Robert C. Jones, Los Altos, CA (US)
Assigned to Fluidigm Corporation, South San Francisco, CA (US)
Appl. No. 14/775,960
Filed by Fluidigm Corporation, South San Francisco, CA (US)
PCT Filed Mar. 14, 2014, PCT No. PCT/US2014/028751
§ 371(c)(1), (2) Date Sep. 14, 2015,
PCT Pub. No. WO2014/144371, PCT Pub. Date Sep. 18, 2014.
Claims priority of provisional application 61/799,559, filed on Mar. 15, 2013.
Prior Publication US 2016/0024557 A1, Jan. 28, 2016
Int. Cl. C12Q 1/68 (2006.01); G01N 33/53 (2006.01); G01N 33/532 (2006.01)
CPC C12Q 1/686 (2013.01) [C12Q 1/682 (2013.01); G01N 33/532 (2013.01); G01N 2458/10 (2013.01)] 11 Claims
 
1. A method of detecting an RNA and a protein in a sample, the method comprising:
(a) incubating a sample in a reaction mixture with a pair of protein-detecting proximity probes comprising a first and a second protein detecting proximity probe where:
the first protein-detecting probe comprises a first antibody that binds to the target protein joined to a first polynucleotide that comprises an I segment at the 3′ end that is complementary to an I segment on the 3′ end of the second probe; and
the second protein-detecting probe comprises a second antibody that binds to the target protein joined to a second polynucleotide that comprises an I segment complementary to the I segment at the 3′ end of the first polynucleotide;
wherein binding of the first antibody to the target protein and binding of the second antibody to the target protein allows the I segment of the first protein proximity probe to hybridize to the I segment of the second protein proximity probe to form a duplex;
(b) extending the duplex using a DNA polymerase to provide an extended product
(c) amplifying the extended product, or a subregion thereof, using a set of primers that amplify the extended product or subregion thereof; and
(d) detecting an RNA of interest in the sample and the amount of amplicon from amplification of the extended product or subregion thereof.