US 12,435,367 B2
Apparatus, system, and method using immiscible-fluid-discrete-volumes
Linda G. Lee, Palo Alto, CA (US); Sam L Woo, Redwood City, VA (US); Congcong Ma, Foster City, CA (US); Richard T. Reel, Hayward, CA (US); Mark F. Oldham, Emerald Hills, CA (US); David M. Cox, Foster City, CA (US); Benjamin G. Schroeder, San Mateo, CA (US); Jon M. Sorenson, Alameda, CA (US); and Willy Wiyatno, Union City, CA (US)
Assigned to APPLIED BIOSYSTEMS, LLC, Carlsbad, CA (US)
Filed by APPLIED BIOSYSTEMS, LLC, Carlsbad, CA (US)
Filed on Oct. 19, 2021, as Appl. No. 17/505,085.
Application 16/054,391 is a division of application No. 14/925,656, filed on Oct. 28, 2015, granted, now 10,041,113, issued on Aug. 7, 2018.
Application 14/925,656 is a division of application No. 12/557,488, filed on Sep. 10, 2009, granted, now 9,194,772, issued on Nov. 24, 2015.
Application 17/505,085 is a continuation of application No. 16/054,391, filed on Aug. 3, 2018, granted, now 11,162,137.
Application 12/557,488 is a continuation of application No. 11/507,735, filed on Aug. 22, 2006, abandoned.
Claims priority of provisional application 60/818,197, filed on Jun. 30, 2006.
Claims priority of provisional application 60/731,133, filed on Oct. 28, 2005.
Claims priority of provisional application 60/710,167, filed on Aug. 22, 2005.
Prior Publication US 2022/0033896 A1, Feb. 3, 2022
Int. Cl. C12Q 1/6869 (2018.01); B01L 3/00 (2006.01); C12Q 1/6806 (2018.01); C12Q 1/6874 (2018.01); F15C 5/00 (2006.01); F16K 99/00 (2006.01); G01N 1/14 (2006.01); G01N 27/447 (2006.01); G01N 35/08 (2006.01); B01L 3/02 (2006.01); B01L 7/00 (2006.01)
CPC C12Q 1/6869 (2013.01) [B01L 3/502715 (2013.01); B01L 3/502784 (2013.01); C12Q 1/6806 (2013.01); C12Q 1/6874 (2013.01); F15C 5/00 (2013.01); F16K 99/0001 (2013.01); F16K 99/0011 (2013.01); F16K 99/0013 (2013.01); G01N 1/14 (2013.01); G01N 27/44743 (2013.01); G01N 27/44769 (2013.01); G01N 35/08 (2013.01); B01J 2219/00353 (2013.01); B01J 2219/0036 (2013.01); B01J 2219/00364 (2013.01); B01J 2219/00608 (2013.01); B01J 2219/0061 (2013.01); B01J 2219/00612 (2013.01); B01J 2219/00619 (2013.01); B01J 2219/00626 (2013.01); B01J 2219/00637 (2013.01); B01J 2219/00653 (2013.01); B01J 2219/00657 (2013.01); B01J 2219/00659 (2013.01); B01L 3/0293 (2013.01); B01L 7/52 (2013.01); B01L 2200/0673 (2013.01); B01L 2200/10 (2013.01); B01L 2300/0864 (2013.01); B01L 2300/0867 (2013.01); B01L 2400/0421 (2013.01); B01L 2400/0487 (2013.01); C12Q 2535/101 (2013.01); F16K 2099/0084 (2013.01); Y10T 137/0318 (2015.04); Y10T 137/4259 (2015.04); Y10T 137/85978 (2015.04); Y10T 137/85986 (2015.04); Y10T 137/86863 (2015.04); Y10T 436/2575 (2015.01)] 20 Claims
 
1. A method comprising:
contacting a stream of aqueous sample fluid flowing in a first conduit with a stream of non-aqueous spacing fluid that is immiscible with the aqueous sample fluid to form discrete volumes of the aqueous sample fluid separated from one another by the non-aqueous spacing fluid, wherein the aqueous sample fluid comprises target nucleic acid, and wherein a first plurality of the discrete volumes contains at least one molecule comprising the target nucleic acid and a second plurality of the discrete volumes contains no molecules comprising the target nucleic acid;
amplifying the target nucleic acid in one or more of the first plurality of the discrete volumes to form an amplicon;
after amplifying the target nucleic acid, adding additional spacing fluid between consecutive discrete volumes of one or both of the first plurality of discrete volumes and the second plurality of discrete volumes, so as to adjust or control spacing between the consecutive discrete volumes;
in a second conduit, detecting a fluorescence signal from the amplicon in the one or more of the first plurality of the discrete volumes; and
based on the detecting, discriminating between the one or more of the first plurality of the discrete volumes and the second plurality of the discrete volumes.