US 12,134,758 B2
Microfluidic cell culture
Randall D. Lowe, Jr., Emeryville, CA (US); Kristin G. Beaumont, New York, NY (US); Aathavan Karunakaran, Berkeley, CA (US); Natalie C. Marks, Albany, CA (US); Jason M. McEwen, El Cerrito, CA (US); Mark P. White, Orinda, CA (US); J. Tanner Nevill, El Cerrito, CA (US); Gang F. Wang, Mountain View, CA (US); Andrew W. McFarland, Berkeley, CA (US); Daniele Malleo, Menlo Park, CA (US); Keith J. Breinlinger, San Rafael, CA (US); Xiao Guan, San Rafael, CA (US); and Kevin T. Chapman, Emeryville, CA (US)
Assigned to BRUKER CELLULAR ANALYSIS, INC., Emeryville, CA (US)
Filed by BRUKER CELLULAR ANALYSIS, INC., Emeryville, CA (US)
Filed on May 12, 2022, as Appl. No. 17/743,318.
Application 17/743,318 is a division of application No. 16/920,365, filed on Jul. 2, 2020, granted, now 11,365,381.
Application 16/920,365 is a continuation of application No. 15/135,707, filed on Apr. 22, 2016, granted, now 10,723,988, issued on Jul. 28, 2020.
Claims priority of provisional application 62/151,325, filed on Apr. 22, 2015.
Prior Publication US 2022/0356429 A1, Nov. 10, 2022
Int. Cl. C12M 3/00 (2006.01); B01L 3/00 (2006.01); C12M 1/00 (2006.01); C12M 3/06 (2006.01)
CPC C12M 23/16 (2013.01) [C12M 29/10 (2013.01)] 20 Claims
OG exemplary drawing
 
1. A method of preparing a microfluidic device comprising:
introducing a conditioning modification reagent comprising a moiety for supporting cell growth, viability, portability, or any combination thereof and a reactive pairing moiety Rpx into a microfluidic circuit of the microfluidic device, the microfluidic device comprising one or more discrete microfluidic circuits each configured to hold a fluid, the microfluidic circuit defined by an enclosure, the enclosure comprising:
a base comprising a first material and having an inner surface;
a microfluidic circuit structure comprising a second material and having an inner surface; and
a cover comprising a third material and having an inner surface, wherein the inner surfaces of the base, microfluidic circuit structure and the cover each face an interior of at least one microfluidic circuit element of the one or more discrete microfluidic circuits,
wherein the inner surfaces of the base, the microfluidic circuit structure and the cover have a monolayer covalently attached thereto, wherein the monolayer comprises a linking group and a reactive moiety Rx configured to covalently link to the conditioning modification reagent within the at least one microfluidic circuit element, and further wherein each linking group of the monolayer of the inner surfaces of the base, the microfluidic circuit structure and the cover is the same linking group; and
covalently linking the reactive pairing moiety Rpx with the reactive moiety Rx thereby forming a conditioned surface monolayer on the inner surfaces of the base, the microfluidic circuit structure and the cover of the at least one microfluidic circuit element, wherein the conditioned surface monolayer is configured to support cell growth, viability, portability, or any combination thereof within the microfluidic device.
 
20. A method for culturing at least one biological cell in a microfluidic device the method comprising:
forming a conditioned surface in a growth chamber of the microfluidic device, the conditioned surface configured to support cell growth, viability, portability, or any combination thereof, wherein forming the conditioned surface comprises introducing a conditioning modification reagent into the microfluidic device, wherein inner surfaces of the microfluidic device have a covalently attached monolayer comprising a linking group and a reactive moiety Rx, wherein the reactive moiety Rx covalently links to the conditioning modification reagent, wherein the conditioning modification reagent has a structure of Formula 5:

OG Complex Work Unit Chemistry
wherein a reactive pairing moiety Rpx is a reactive group that reacts with the reactive moiety Rx;
moiety is a moiety configured to support cell growth, viability, portability or any combination thereof;
L′ is a linker having 1 to 100 non-hydrogen atoms selected from any combination of silicon, carbon, nitrogen, oxygen, sulfur and phosphorus atoms; and
m is 0 or 1;
introducing the at least one biological cell into the growth chamber having the conditioned surface configured to support cell growth, viability, portability, or any combination thereof,
wherein the microfluidic device comprises a discrete microfluidic circuit configured to contain a flow of a first fluidic medium, the discrete microfluidic circuit comprising a fluid inlet port, the growth chamber, and a flow region connecting the fluid inlet port with the growth chamber;
incubating the at least one biological cell in the microfluidic device for a period of time long enough to expand the at least one biological cell to produce a colony of biological cells; and
non-continuously perfusing the first fluidic medium through the flow region during the incubating step.