High-throughput single-cell imaging, sorting, and isolation
Inventors
Bharadwaj, Rajiv • Fathollahi, Bahram
Assignees
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Publication Number
US-9165359-B2
Publication Date
2015-10-20
Expiration Date
Abstract
The invention provides a method and apparatus for isolating individual target cells. The apparatus includes a body structure comprising a main channel, a collection channel, and a waste channel fluidly coupled at a first fluid junction. A plurality of trapping channels intersect the collection channel, each trapping channel having a diameter at a location adjacent to the intersection of the trapping channel with the collection channel that is less than a diameter of an individual target cell. The apparatus also includes an imaging system configured to image individual target and non-target cells within the main channel, thereby producing imaging data; a processor configured to perform real-time, multivariate analyses of the imaging data; and a directing system configured to direct the individual target cells. A pressure source is in fluid communication with one or more of the collection channel, the waste channel, the first side channel, and the second side channel.
Core Innovation
The invention provides an apparatus for isolating individual target cells with a microfluidic body structure that includes a main channel, a collection channel, and a waste channel fluidly coupled at a first fluid junction. The body structure further includes a plurality of trapping channels intersecting the collection channel, where each trapping channel has a diameter at a location adjacent to the intersection that is less than a diameter of an individual target cell.
A focusing system includes first and second focusing fluid channels intersecting the main channel at a second fluid junction on opposite sides of the main channel, producing a flow of individual target and non-target cells within the main channel. The apparatus includes an imaging system configured to image each of a plurality of individual target and non-target cells within the main channel to produce imaging data selected from the group consisting of high-resolution imaging data and high-content imaging data. A processor performs real-time multivariate analyses of the imaging data, and a directing system directs individual target cells.
The invention also provides a method for isolating individual target cells using the same microfluidic body structure and processing chain. Target and non-target cells are flowed through the main channel, imaging data are obtained in an imaging area, and a processor performs real-time multivariate analysis to identify each cell as a target cell or a non-target cell. When a cell is identified as a target cell, the target cell is directed into the collection channel, immobilized at a trapping location using a first pressure, and then drawn into and through the trapping channel at the trapping location using a second pressure.
Claims Coverage
Two independent claims are present: an apparatus claim and a method claim. Each independent claim centers on microfluidic trapping-channel sizing for individual target cells, imaging with high-resolution or high-content imaging data, real-time multivariate analysis, and pressure-controlled immobilization and drawing at trapping locations, with routing of target versus non-target cells to different channels.
Microfluidic trapping channels sized smaller than individual target cells
A body structure comprising a main channel, a collection channel, and a waste channel fluidly coupled at a first fluid junction, with a plurality of trapping channels intersecting the collection channel, each trapping channel having a diameter at a location adjacent to the intersection that is less than a diameter of an individual target cell.
Opposed-side focusing fluid channels for individual cell imaging in the main channel
A first focusing fluid channel intersecting the main channel at a second fluid junction and a second focusing fluid channel intersecting the main channel at the second fluid junction on opposite sides of the main channel, with a focusing system comprising a first focusing fluid in fluid communication with the first focusing fluid channel and a second focusing fluid in fluid communication with the second focusing fluid channel.
Imaging high-resolution or high-content imaging data with processor multivariate analysis
An imaging system configured to image each of a plurality of individual target and non-target cells within the main channel to produce imaging data selected from the group consisting of high-resolution imaging data and high-content imaging data, and a processor configured to perform real-time, multivariate analyses of the imaging data.
Directing system routes individual target cells with pressure source control
A directing system configured to direct the individual target cells, and a pressure source in fluid communication with one or more of the collection channel, the waste channel, the first side channel, and the second side channel.
Real-time classification and routing to collection or waste channels
Providing a body structure comprising a main channel, a collection channel, and a waste channel fluidly coupled at a first fluid junction with a plurality of trapping channels intersecting the collection channel with trapping locations adjacent to the intersection, each trapping location having a diameter less than a diameter of an individual target cell; flowing a plurality of target and non-target cells through the main channel; obtaining imaging data selected from the group consisting of high-resolution imaging data and high-content imaging data; using a processor to perform a real-time, multivariate analysis to identify the cell as a target cell or a non-target cell; and, when identified as a target cell, directing the target cell into the collection channel.
Immobilize at trapping location with first pressure and draw through trapping channel with second pressure
Applying a first pressure to immobilize the target cell at the trapping location of one of the plurality of trapping channels, repeating steps c through f for each target and non-target cell until all of the target cells flowing through the main channel have been immobilized or until a target cell has been immobilized at the trapping location of all of the plurality of trapping channels, whichever event occurs first, and applying a second pressure to draw one or more of the target cells into and through the trapping channel at whose trapping location the target cell has been immobilized.
Overall, the independent claims combine (i) microfluidic trapping channels with diameters less than the target cell diameter at trapping locations, (ii) imaging producing high-resolution or high-content imaging data, (iii) real-time multivariate analysis to classify target versus non-target cells, (iv) directing of target cells into a collection channel, and (v) pressure-controlled immobilization at trapping locations followed by drawing through the trapping channel.
Stated Advantages
Not explicitly described in patent.
Documented Applications
Not explicitly described in patent.
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