Instruments, modules, and methods for improved detection of edited sequences in live cells
First Claim
Patent Images
1. A singulation assembly for a solid wall singulation or substantial singulation, growth, induction of editing, and normalization or cherry-picking (SWIIN) module comprising:
- a retentate member comprising an upper surface and a lower surface, wherein the retentate member comprises at least one retentate distribution channel which traverses the retentate member from its upper surface to its lower surface and for 70% to 95% of a length of retentate member;
wherein the lower surface of the retentate member comprises flow directors;
wherein the retentate member further comprises one or more retentate member ports configured to supply cells and fluid to and remove cells and fluid from the retentate member; and
wherein the retentate member ports are fluidically-connected to the at least one retentate distribution channel and retentate flow directors;
a perforated member with an upper surface and a lower surface, wherein the upper surface of the perforated member is positioned beneath and adjacent to the lower surface of the retentate member and wherein the perforated member comprises at least 25,000 perforations;
a filter with an upper surface and a lower surface, wherein the upper surface of the filter is positioned beneath and adjacent to the lower surface of the perforated member and wherein the lower surface of the filter is positioned above and adjacent to an upper surface of a permeate member; and
the permeate member comprising the upper surface and a lower surface, wherein the permeate member comprises at least one permeate distribution channel which traverses the permeate member from its lower surface to its upper surface and for 70% to 95% of a length of permeate member;
wherein the upper surface of the permeate member comprises permeate flow directors;
wherein the permeate member further comprises one or more permeate member ports configured to supply fluid to and remove fluid from the permeate member; and
wherein the permeate member ports are fluidically-connected to the at least one permeate distribution channel and permeate flow directors; and
adhesive to couple the retentate member, perforated member, filter and permeate member.
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Abstract
The present disclosure provides instruments, modules and methods for improved detection of edited cells following nucleic acid-guided nuclease genome editing. The disclosure provides improved automated instruments that perform methods—including high throughput methods—for screening cells that have been subjected to editing and identifying cells that have been properly edited.
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Citations
20 Claims
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1. A singulation assembly for a solid wall singulation or substantial singulation, growth, induction of editing, and normalization or cherry-picking (SWIIN) module comprising:
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a retentate member comprising an upper surface and a lower surface, wherein the retentate member comprises at least one retentate distribution channel which traverses the retentate member from its upper surface to its lower surface and for 70% to 95% of a length of retentate member;
wherein the lower surface of the retentate member comprises flow directors;
wherein the retentate member further comprises one or more retentate member ports configured to supply cells and fluid to and remove cells and fluid from the retentate member; and
wherein the retentate member ports are fluidically-connected to the at least one retentate distribution channel and retentate flow directors;a perforated member with an upper surface and a lower surface, wherein the upper surface of the perforated member is positioned beneath and adjacent to the lower surface of the retentate member and wherein the perforated member comprises at least 25,000 perforations; a filter with an upper surface and a lower surface, wherein the upper surface of the filter is positioned beneath and adjacent to the lower surface of the perforated member and wherein the lower surface of the filter is positioned above and adjacent to an upper surface of a permeate member; and the permeate member comprising the upper surface and a lower surface, wherein the permeate member comprises at least one permeate distribution channel which traverses the permeate member from its lower surface to its upper surface and for 70% to 95% of a length of permeate member;
wherein the upper surface of the permeate member comprises permeate flow directors;
wherein the permeate member further comprises one or more permeate member ports configured to supply fluid to and remove fluid from the permeate member; and
wherein the permeate member ports are fluidically-connected to the at least one permeate distribution channel and permeate flow directors; andadhesive to couple the retentate member, perforated member, filter and permeate member. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15)
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16. A SWIIN module comprising:
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a singulation assembly comprising; a retentate member comprising an upper surface and a lower surface, wherein the retentate member comprises at least one retentate distribution channel which traverses the retentate member from its upper surface to its lower surface and for 70% to 95% of a length of retentate member;
wherein the lower surface of the retentate member comprises retentate flow directors;
wherein the one or more retentate member further comprises one or more retentate member ports configured to supply cells and fluid to and remove cells and fluid from the retentate member; and
wherein the retentate member ports are fluidically-connected to the at least one retentate distribution channel and retentate flow directors;a perforated member with an upper surface and a lower surface, wherein the upper surface of the perforated member is positioned beneath and adjacent to the lower surface of the retentate member; a filter with an upper surface and a lower surface, wherein the upper surface of the filter is positioned beneath and adjacent to the lower surface of the perforated member and wherein the lower surface of the filter is positioned above and adjacent to an upper surface of a permeate member; and the permeate member comprising the upper surface and a lower surface, wherein the permeate member comprises at least one permeate distribution channel which traverses the permeate member from its lower surface to its upper surface and for 70% to 95% of a length of permeate member;
wherein the upper surface of the permeate member comprises permeate flow directors;
wherein the permeate member further comprises one or more permeate member ports configured to supply fluid to and remove fluid from the permeate member; and
wherein the one or more permeate member ports are fluidically-connected to the at least one permeate distribution channel and permeate flow directors; andadhesive to couple the retentate member, perforated member, filter and permeate member; a reservoir assembly comprising at least two reservoirs wherein a first reservoir is
1) fluidically-coupled to a first reservoir aperture into which fluids and/or cells flow from outside the SWIIN module into the first reservoir,
2) fluidically-coupled to at least one reservoir/channel port from which fluids and/or cells flow from the first reservoir into the one or more retentate member ports; and
3) pneumatically-coupled via a first pneumatic access aperture to a pressure source;a second reservoir is
1) fluidically-coupled to a second reservoir access aperture into which fluids and/or cells flow from outside the SWIIN module into the second reservoir,
2) fluidically-coupled to a reservoir/channel port from which fluids and/or cells flow from the second reservoir into the one or more permeate member ports; and
3) pneumatically-coupled via a second pneumatic access aperture to a pressure source; anda SWIIN cover. - View Dependent Claims (17, 18, 19, 20)
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Specification