US 2022/0010360 A1Published application

Multiomic analysis of cell analytes using microfluidic systems

AssigneeCornell University

InventorsHarold G. CRAIGHEAD, Harvey C. TIAN, David M. LIN, Adam J. BISOGNI

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Abstract

A method for isolating one or more distinct analyte component from a cell sample is disclosed, as well as processes for testing and analyzing the distinct analyte components. The distinct analyte components include: (i) a total protein fraction; (ii) a plasma membrane protein fraction; (iii) a total RNA fraction; (iv) a cytosolic RNA fraction; (v) a cytosolic protein fraction; (vi) a nuclear RNA fraction; (vii) a nuclear protein fraction; (viii) a chromatin fraction comprising genomic DNA (gDNA); (ix) a gDNA markers fraction. This method involves the use of microfluidic device having a cell capture component and a nucleic acid entanglement component, where the cell capture component includes a cell capture array having a plurality of cell capture micropillars, where the nucleic acid entanglement component includes a nucleic acid entanglement array having a plurality of nucleic acid entanglement micropillars, and where the microfluidic device operates under continuous flow conditions.

Claims50

Dependent claims

A method for isolating one or more distinct analyte component from a cell sample, said method comprising the steps of:

(a) providing a microfluidic device comprising a cell capture component and a nucleic acid entanglement component, wherein said cell capture component comprises a cell capture array comprising a plurality of cell capture micropillars, wherein said nucleic acid entanglement component comprises a nucleic acid entanglement array comprising a plurality of nucleic acid entanglement micropillars, and wherein the microfluidic device operates under continuous flow conditions;
(b) capturing one or more cell in the cell capture array of the microfluidic device while being subjected to a continuous flow rate, said capturing step comprising introducing the one or more cell sample into the microfluidic device under sufficient hydrodynamic flow to entrap, by size exclusion, the one or more cell within the cell capture component;
(c) treating the one or more captured cell with a sequential workflow procedure under conditions effective to separate one or more distinct analyte component therefrom, wherein said one or more distinct analyte component is selected from the group consisting of:
(i) a total protein fraction;
(ii) a plasma membrane protein fraction;
(iii) a total RNA fraction;
(iv) a cytosolic RNA fraction;
(v) a cytosolic protein fraction;
(vi) a nuclear RNA fraction;
(vii) a nuclear protein fraction;
(viii) a chromatin fraction comprising genomic DNA (gDNA) regions of open chromatin;
(ix) a gDNA markers fraction comprising epigenetic and regulatory markers of gDNA;
(x) an amplified gDNA fraction;
and
(xi) a methylated gDNA fraction;
and
(d) isolating the one or more distinct analyte component in a manner suitable for further testing and/or analysis thereof.

14 more elements

Dependencies are inferred from the claim text.

Facts

Application: 17/298,533Filed 29 Nov 2019
Priority: 28 Nov 2018
Published: 13 Jan 2022
Term ends: Not estimatedThe source records don’t support an estimate.

Classification12 codes

C12Q 1/6806
Involving nucleic acids: preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay
B01L 3/5027
With fluid transport, e.g. in multi-compartment structures: by integrated microfluidic structures, i.e. dimensions of channels and chambers are…
B01L 3/502761
By integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g.…
C12N 15/1006
Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses…