Massive Parallel Analysis of Single Cells in an Integrated Microfluidic Platform

Jimenez-Valdes, Rocio J.; Rodriguez‐Moncayo, Roberto; Cedillo-Alcantar, Diana F.; García-Cordero, José L.

doi:10.1021/acs.analchem.6b04485

Cited by 20 publications

(17 citation statements)

References 48 publications

(68 reference statements)

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“…To demonstrate the utility of our microscope in quantitative biological experiments, we performed a fluorescence time-lapse experiment to track the production of neutrophil extracellular traps (NETs) from single cells [26]. The nuclei of neutrophils isolated from peripheral blood were first stained with Hoechst, and then captured in a PDMS device containing an array of microwells (20 μm in diameter).…”

Section: Resultsmentioning

confidence: 99%

Facile assembly of an affordable miniature multicolor fluorescence microscope made of 3D-printed parts enables detection of single cells

Tristan-Landin¹,

Gonzalez-Suarez²,

Jimenez-Valdes³

et al. 2019

Preprint

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18Fluorescence microscopy is one of the workhorses of biomedical research and laboratory diagnosis;19 however, their cost, size, maintenance, and fragility has prevented their adoption in developing countries 20 or low-resource settings. Although significant advances have decreased their size, cost and accessibility, 21 their designs and assembly remain rather complex. Here, inspired on the simple mechanism from a nut and 22 a bolt we report the construction of a portable fluorescence microscope that operates in bright field mode 23 and in three fluorescence channels: UV, green, and red. It is assembled in under 10 min from only six 3D 24printed parts and basic electronic components that can be readily purchased in most locations or online for 25 US $85. Adapting a microcomputer and a touch LCD screen, the microscope can capture time-lapse images 26 and videos. We characterized its resolution and illumination conditions and benchmarked its performance 27 against a high-end fluorescence microscope by tracking a biological process in single cells. We also 28 demonstrate its application to image cells inside a microfluidic device in bright-field and fluorescence 29 mode. Our microscope fits in a CO 2 chamber and can be operated in time-lapse mode. Our portable 30 microscope is ideal in applications where space is at a premium, such as lab-on-a-chips or space missions, 31and can find applications in clinical research, diagnostics, telemedicine and in educational settings. 32 33 3 34

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Section: Resultsmentioning

confidence: 99%

Facile assembly of an affordable miniature multicolor fluorescence microscope made of 3D-printed parts enables detection of single cells

Tristan-Landin¹,

Gonzalez-Suarez²,

Jimenez-Valdes³

et al. 2019

Preprint

Self Cite

View full text Add to dashboard Cite

show abstract

“…To demonstrate the utility of our microscope in quantitative biological experiments, we performed a fluorescence time-lapse experiment to track the production of neutrophil extracellular traps (NETs) from single cells [29]. The nuclei of neutrophils isolated from peripheral blood were first stained with Hoechst, and then captured in a PDMS device containing an array of microwells (20 μm in diameter and depth).…”

Section: Resultsmentioning

confidence: 99%

Facile assembly of an affordable miniature multicolor fluorescence microscope made of 3D-printed parts enables detection of single cells

et al. 2019

Self Cite

View full text Add to dashboard Cite

Fluorescence microscopy is one of the workhorses of biomedical research and laboratory diagnosis; however, their cost, size, maintenance, and fragility has prevented their adoption in developing countries or low-resource settings. Although significant advances have decreased their size, cost and accessibility, their designs and assembly remain rather complex. Here, inspired on the simple mechanism from a nut and a bolt, we report the construction of a portable fluorescence microscope that operates in bright-field mode and in three fluorescence channels: UV, green, and red. It is assembled in under 10 min from only six 3D printed parts, basic electronic components, a microcomputer (Raspberry Pi) and a camera, all of which can be readily purchased in most locations or online for US $122. The microcomputer was programmed in Python language to capture time-lapse images and videos. Resolution and illumination conditions of the microscope were characterized, and its performance was compared with a high-end fluorescence microscope in bright-field and fluorescence mode. We demonstrate that our miniature microscope can resolve and track single cells in both modes. The instructions on how to assemble the microscope are shown in a video, and the software to control it and the design files of the 3D-printed parts are freely available online. Our portable microscope is ideal in applications where space is at a premium, such as lab-on-a-chips or space missions, and can find applications in basic and clinical research, diagnostics, telemedicine and in educational settings.

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“…Each of the nine outlets from the CGG leads to a single microchamber, where cells are exposed in parallel to different concentrations of the same molecule. To capture cells, we employ microwells, a technology that allows cells to experience low shear-stresses while being perfused at different flow rates . After the chemical stimuli passes through all the microchambers they converge into a single outlet for waste removal.…”

Section: Resultsmentioning

confidence: 99%

Dynamic Generation of Concentration- and Temporal-Dependent Chemical Signals in an Integrated Microfluidic Device for Single-Cell Analysis

et al. 2018

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Intracellular signaling pathways are affected by the temporal nature of external chemical signaling molecules such as neurotransmitters or hormones. Developing high-throughput technologies to mimic these time-varying chemical signals and to analyze the response of single cells would deepen our understanding of signaling networks. In this work, we introduce a microfluidic platform to stimulate hundreds of single cells with chemical waveforms of tunable frequency and amplitude. Our device produces a linear gradient of 9 concentrations that are delivered to an equal number of chambers, each containing 492 microwells, where individual cells are captured. The device can alternate between the different stimuli concentrations and a control buffer, with a maximum operating frequency of 33 mHz that can be adjusted from a computer. Fluorescent time-lapse microscopy enables to obtain hundreds of thousands of data points from one experiment. We characterized the gradient performance and stability by staining hundreds of cells with calcein AM. We also assessed the capacity of our device to introduce periodic chemical stimuli of different amplitudes and frequencies. To demonstrate our device performance, we studied the dynamics of intracellular Ca release from intracellular stores of HEK cells when stimulated with carbachol at 4.5 and 20 mHz. Our work opens the possibility of characterizing the dynamic responses in real time of signaling molecules to time-varying chemical stimuli with single cell resolution.

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Massive Parallel Analysis of Single Cells in an Integrated Microfluidic Platform

Cited by 20 publications

References 48 publications

Facile assembly of an affordable miniature multicolor fluorescence microscope made of 3D-printed parts enables detection of single cells

Facile assembly of an affordable miniature multicolor fluorescence microscope made of 3D-printed parts enables detection of single cells

Facile assembly of an affordable miniature multicolor fluorescence microscope made of 3D-printed parts enables detection of single cells

Dynamic Generation of Concentration- and Temporal-Dependent Chemical Signals in an Integrated Microfluidic Device for Single-Cell Analysis

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