A fast cell loading and high‐throughput microfluidic system for long‐term cell culture in zero‐flow environments

Luo, Chunxiong; Zhu, Xuejun; Yu, Tao; Luo, Xianjia; Ouyang, Qi; Ji, Hang; Chen, Yong

doi:10.1002/bit.21877

Cited by 54 publications

(43 citation statements)

References 21 publications

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“…Various microfluidic devices maintain cells in a single focal plane as they grow (10,(16)(17)(18)(19)(20)(21), but many of these devices require sophisticated fabrication techniques such as multilayer fabrication with valves (16,18), channel height differences (17), or membranes (10,21). To optimize the statistical power of these techniques, the initial placement of cells should be controlled; several other microfluidic devices achieve single-cell trapping (22)(23)(24), but these trapping mechanisms are not conducive to the lineage analysis that we perform here. The ability to robustly and repeatedly trap, spatially organize, and track the growth of single cells over many generations in a device that is easy to fabricate and simple to use would enable the collection of data over many cell lineages in a single experiment.…”

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confidence: 99%

Tracking lineages of single cells in lines using a microfluidic device

Rowat

Bird²,

Agresti³

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

114

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Cells within a genetically identical population exhibit phenotypic variation that in some cases can persist across multiple generations. However, information about the temporal variation and familial dependence of protein levels remains hidden when studying the population as an ensemble. To correlate phenotypes with the age and genealogy of single cells over time, we developed a microfluidic device that enables us to track multiple lineages in parallel by trapping single cells and constraining them to grow in lines for as many as 8 divisions. To illustrate the utility of this method, we investigate lineages of cells expressing one of 3 naturally regulated proteins, each with a different representative expression behavior. Within lineages deriving from single cells, we observe genealogically related clusters of cells with similar phenotype; cluster sizes vary markedly among the 3 proteins, suggesting that the time scale of phenotypic persistence is protein-specific. Growing lines of cells also allows us to dynamically track temporal fluctuations in protein levels at the same time as pedigree relationships among the cells as they divide in the chambers. We observe bursts in expression levels of the heat shock protein Hsp12-GFP that occur simultaneously in mother and daughter cells. In contrast, the ribosomal protein Rps8b-GFP shows relatively constant levels of expression over time. This method is an essential step toward understanding the time scales of phenotypic variation and correlations in phenotype among single cells within a population.epigenetics ͉ protein expression ͉ single cell assay ͉ yeast

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mentioning

confidence: 99%

Tracking lineages of single cells in lines using a microfluidic device

Rowat

Bird²,

Agresti³

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

114

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“…Due to gas absorption by the PDMS surface, the cell suspension is passively aspirated through the cell migration channel and into micro-cavities. 40 After approximately 10 min, cells had gathered within the cell-loading cavity, and both cell-loading cavity and liquid-absorbing cavity were fully filled by cell medium solution. Then, the microfluidic chip was returned to the incubator (37 C, 5% CO 2 ) for 4 h to allow cells to adhere to its surface.…”

Section: Cell Migration In Different Oxygen Environmentsmentioning

confidence: 99%

A novel microfluidic platform for studying mammalian cell chemotaxis in different oxygen environments under zero-flow conditions

et al. 2015

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The cell's micro-environment plays an important role in various physiological and pathological phenomena. To better investigate in vivo cellular behaviors, researchers have expended great effort in building controlled in vitro biophysical and biochemical environments. Because a cell's gaseous environment affects properties such as its division, metastasis, and differentiation, we developed a zero-flow based platform for studying mammalian cell chemotaxis behavior in different oxygen environments. This platform can construct a linear range of oxygen tensions within one chip (i.e., from 1.4% to 3.6% or 5.5% to 14.5%). To study cell chemotaxis behavior under varying oxygen environments, the chemical gradient direction is established perpendicularly to oxygen change within an observation area. Because the observation area is not subject to flow, shear force is of no concern. In addition, water flow around the cell chambers greatly reduces evaporation and makes long-term microscope imaging possible. In this study, we precisely measure the chemotaxis velocity of MCF-7 human breast cancer cells under different oxygen tension conditions towards CXCL12, which is a stromal cell-derived factor. We find that cell migration rates are not equivalent, even under two close oxygen tensions. We also observed that cells move faster towards high concentrations of chemoattractant when the oxygen tension is below 3% due to the increased expression of HIF-1 (hypoxia-inducible factor 1), which promotes a transition to the amoeboid rather than mesenchymal mode of movement. Our experiments demonstrate that this new microfluidic platform is useful for the quantitative study of mammalian cell chemotaxis under different oxygen conditions in the absence of shear force. We also shed light on the study of chemotaxis under other gaseous environments. V C 2015 AIP Publishing LLC.

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“…First, PDMS molds were degassed in vacuum for 30 min (Luo et al, 2008) and then placed on the surface of a glass or PS Petri dish substrate. Afterward, the prepared solution was added in the capillary entrance of the PDMS mold, as shown in Figure 1.…”

Section: Degassing-assisted Patterningmentioning

confidence: 99%

“…When a PDMS stamp is degassed in a rough vacuum and then brought back to atmosphere, it will reabsorb air toward a new equilibrium (Merkel et al, 2000). Therefore, a degassed PDMS mold placed on a flat surface substrate can be used for pump-free manipulation of any microfluidic samples (Hosokawa et al, 2004(Hosokawa et al, , 2006Luo et al, 2008;Zhou et al, 2007). After drying or incubation, a variety of surface patterns can be thus produced without applying external pressure.…”

mentioning

confidence: 98%

Degassing‐assisted patterning of cell culture surfaces

Luo

Liu

et al. 2009

Biotech & Bioengineering

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We developed an alternative patterning technique which is capable of producing both topographic and biochemical features for cell culture studies. This technique is based on microaspiration induced with a degassed poly (dimethylsiloxane) (PDMS) mold. After degassing in a rough vacuum chamber and placed on a sample surface, liquid solution can be aspired through channels and cavities created in the PDMS mold. Depending on the composition of the solution and the associated drying or incubation processes, a variety of surface patterns can be produced without applying external pressure. For demonstration, we fabricated agarose gel microwells and biomolecule patterns either on a glass plate or in a cell culture Petri dish, both applicable for cell culture studies.

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A fast cell loading and high‐throughput microfluidic system for long‐term cell culture in zero‐flow environments

Cited by 54 publications

References 21 publications

Tracking lineages of single cells in lines using a microfluidic device

Tracking lineages of single cells in lines using a microfluidic device

A novel microfluidic platform for studying mammalian cell chemotaxis in different oxygen environments under zero-flow conditions

Degassing‐assisted patterning of cell culture surfaces

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