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High-Content Quantification of Single-Cell Immune Dynamics.

Junkin M, Kaestli AJ, Cheng Z, Jordi C, Albayrak C, Hoffmann A, Tay S - Cell Rep (2016)

Bottom Line: Characterizing dynamic input-output relationships in single cells is crucial for understanding and modeling cellular systems.We developed an automated microfluidic system that delivers precisely defined dynamical inputs to individual living cells and simultaneously measures key immune parameters dynamically.Our system combines nanoliter immunoassays, microfluidic input generation, and time-lapse microscopy, enabling study of previously untestable aspects of immunity by measuring time-dependent cytokine secretion and transcription factor activity from single cells stimulated with dynamic inflammatory inputs.

View Article: PubMed Central - PubMed

Affiliation: Department of Biosystems Science and Engineering, ETH Zürich, 4058 Basel, Switzerland.

No MeSH data available.


Related in: MedlinePlus

Automated Microfluidic System for Multiparameter Analysis of Single-Cell Immune Dynamics(A) Top left: microfluidic device with functional areas indicated. The scale bar represents 5 mm. Middle right: close-up of a single functional column. The arrows show medium movement during sampling (solid) and circulation for mixing during binding (dashed). Right: close-ups showing a cell captured inside a culture chamber (red), a binding chamber (blue) with a retained bead, and beads inside storage chambers (red). Bottom left: fluorescent imaging of NF-κB activation (top row). The arrow shows the activated cell (RelA in nucleus), and the scale bar represents 20 μm. Middle row: phase contrast images. Bottom row: brightfield-fluorescence merged images of beads. The red spot in the third panel indicates fluorescent sandwich immune assay detection of secreted protein.(B) Setup of microfluidic device mounted on automated microscope inside atmospheric enclosure and image of control software interface.(C) Operation of secretion assay. Beads are initially loaded into storage chambers and then moved by direction of control software to binding chamber. There they are exposed to medium coming from cells through peristaltic pumping. Once analyte is mixed and bound, beads are rinsed and moved back to original storage chambers. This process is repeated with a new bead for different time points so that each row corresponds to a specific time point.See also Figure S1.
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fig1: Automated Microfluidic System for Multiparameter Analysis of Single-Cell Immune Dynamics(A) Top left: microfluidic device with functional areas indicated. The scale bar represents 5 mm. Middle right: close-up of a single functional column. The arrows show medium movement during sampling (solid) and circulation for mixing during binding (dashed). Right: close-ups showing a cell captured inside a culture chamber (red), a binding chamber (blue) with a retained bead, and beads inside storage chambers (red). Bottom left: fluorescent imaging of NF-κB activation (top row). The arrow shows the activated cell (RelA in nucleus), and the scale bar represents 20 μm. Middle row: phase contrast images. Bottom row: brightfield-fluorescence merged images of beads. The red spot in the third panel indicates fluorescent sandwich immune assay detection of secreted protein.(B) Setup of microfluidic device mounted on automated microscope inside atmospheric enclosure and image of control software interface.(C) Operation of secretion assay. Beads are initially loaded into storage chambers and then moved by direction of control software to binding chamber. There they are exposed to medium coming from cells through peristaltic pumping. Once analyte is mixed and bound, beads are rinsed and moved back to original storage chambers. This process is repeated with a new bead for different time points so that each row corresponds to a specific time point.See also Figure S1.

Mentions: A two-layer polydimethylsiloxane (PDMS) microfluidic chip (Melin and Quake, 2007, Kellogg et al., 2014) was designed (Figures 1 and S1), which cultures single cells inside 40 isolated 1.35-nl chambers under the control of an automated culture and measurement system. The device utilizes thousands of PDMS membrane valves to trap and maintain single live cells inside completely isolated chambers while providing dynamic stimulations and non-destructively measuring parameters including transcription factor activation, cytokine secretion, position, and morphology. The microfluidic device is mounted on an automated stage of an inverted epifluorescence microscope placed inside an enclosure, which controls temperature, humidity, and gas composition for mammalian cell culture (Figure 1B).


High-Content Quantification of Single-Cell Immune Dynamics.

Junkin M, Kaestli AJ, Cheng Z, Jordi C, Albayrak C, Hoffmann A, Tay S - Cell Rep (2016)

Automated Microfluidic System for Multiparameter Analysis of Single-Cell Immune Dynamics(A) Top left: microfluidic device with functional areas indicated. The scale bar represents 5 mm. Middle right: close-up of a single functional column. The arrows show medium movement during sampling (solid) and circulation for mixing during binding (dashed). Right: close-ups showing a cell captured inside a culture chamber (red), a binding chamber (blue) with a retained bead, and beads inside storage chambers (red). Bottom left: fluorescent imaging of NF-κB activation (top row). The arrow shows the activated cell (RelA in nucleus), and the scale bar represents 20 μm. Middle row: phase contrast images. Bottom row: brightfield-fluorescence merged images of beads. The red spot in the third panel indicates fluorescent sandwich immune assay detection of secreted protein.(B) Setup of microfluidic device mounted on automated microscope inside atmospheric enclosure and image of control software interface.(C) Operation of secretion assay. Beads are initially loaded into storage chambers and then moved by direction of control software to binding chamber. There they are exposed to medium coming from cells through peristaltic pumping. Once analyte is mixed and bound, beads are rinsed and moved back to original storage chambers. This process is repeated with a new bead for different time points so that each row corresponds to a specific time point.See also Figure S1.
© Copyright Policy - CC BY-NC-ND
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4835544&req=5

fig1: Automated Microfluidic System for Multiparameter Analysis of Single-Cell Immune Dynamics(A) Top left: microfluidic device with functional areas indicated. The scale bar represents 5 mm. Middle right: close-up of a single functional column. The arrows show medium movement during sampling (solid) and circulation for mixing during binding (dashed). Right: close-ups showing a cell captured inside a culture chamber (red), a binding chamber (blue) with a retained bead, and beads inside storage chambers (red). Bottom left: fluorescent imaging of NF-κB activation (top row). The arrow shows the activated cell (RelA in nucleus), and the scale bar represents 20 μm. Middle row: phase contrast images. Bottom row: brightfield-fluorescence merged images of beads. The red spot in the third panel indicates fluorescent sandwich immune assay detection of secreted protein.(B) Setup of microfluidic device mounted on automated microscope inside atmospheric enclosure and image of control software interface.(C) Operation of secretion assay. Beads are initially loaded into storage chambers and then moved by direction of control software to binding chamber. There they are exposed to medium coming from cells through peristaltic pumping. Once analyte is mixed and bound, beads are rinsed and moved back to original storage chambers. This process is repeated with a new bead for different time points so that each row corresponds to a specific time point.See also Figure S1.
Mentions: A two-layer polydimethylsiloxane (PDMS) microfluidic chip (Melin and Quake, 2007, Kellogg et al., 2014) was designed (Figures 1 and S1), which cultures single cells inside 40 isolated 1.35-nl chambers under the control of an automated culture and measurement system. The device utilizes thousands of PDMS membrane valves to trap and maintain single live cells inside completely isolated chambers while providing dynamic stimulations and non-destructively measuring parameters including transcription factor activation, cytokine secretion, position, and morphology. The microfluidic device is mounted on an automated stage of an inverted epifluorescence microscope placed inside an enclosure, which controls temperature, humidity, and gas composition for mammalian cell culture (Figure 1B).

Bottom Line: Characterizing dynamic input-output relationships in single cells is crucial for understanding and modeling cellular systems.We developed an automated microfluidic system that delivers precisely defined dynamical inputs to individual living cells and simultaneously measures key immune parameters dynamically.Our system combines nanoliter immunoassays, microfluidic input generation, and time-lapse microscopy, enabling study of previously untestable aspects of immunity by measuring time-dependent cytokine secretion and transcription factor activity from single cells stimulated with dynamic inflammatory inputs.

View Article: PubMed Central - PubMed

Affiliation: Department of Biosystems Science and Engineering, ETH Zürich, 4058 Basel, Switzerland.

No MeSH data available.


Related in: MedlinePlus