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Biomicrofluidics / Volume 2 / Issue 2 / REGULAR ARTICLES

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Biomicrofluidics 2, 024105 (2008); doi:10.1063/1.2952290 (12 pages)

A transparent cell-culture microchamber with a variably controlled concentration gradient generator and flow field rectifier

Ji-Yen Cheng1, Meng-Hua Yen1,2, Ching-Te Kuo1, and Tai-Horng Young2

1Research Center for Applied Sciences, Academia Sinica, Taiwan 11529
2Institute of Biomedical Engineering, National Taiwan University, Taiwan 10051

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(Received 13 February 2008; accepted 3 June 2008; published online 17 June 2008)

Real-time observation of cell growth provides essential information for studies such as cell migration and chemotaxis. A conventional cell incubation device is usually too clumsy for these applications. Here we report a transparent microfluidic device that has an integrated heater and a concentration gradient generator. A piece of indium tin oxide (ITO) coated glass was ablated by our newly developed visible laser-induced backside wet etching (LIBWE) so that transparent heater strips were prepared on the glass substrate. A polymethylmethacrylate (PMMA) microfluidic chamber with flow field rectifiers and a reagent effusion hole was fabricated by a CO2 laser and then assembled with the ITO heater so that the chamber temperature can be controlled for cell culturing. A variable chemical gradient was generated inside the chamber by combining the lateral medium flow and the flow from the effusion hole. Successful culturing was performed inside the device. Continuous long-term (>10 days) observation on cell growth was achieved. In this work the flow field, medium replacement, and chemical gradient in the microchamber are elaborated.

© 2008 American Institute of Physics

Article Outline

  1. INTRODUCTION
  2. Experimental
    1. Chip design and fabrication
    2. Flow field and chemical gradient observation
    3. Flow simulation
    4. Temperature distribution measurement
    5. Cell culture in microchamber
  3. RESULT AND DISCUSSION
    1. Medium flow and replacement
    2. Chemical gradient buildup and control
    3. Temperature homogeneity and cell culture
  4. CONCLUSION

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KEYWORDS and PACS

Keywords

bioMEMS, cellular biophysics, laser beam applications, microfluidics

PACS

  • 87.80.Ek

    Mechanical and micromechanical techniques

  • 07.10.Cm

    Micromechanical devices and systems

ARTICLE DATA

Permalink
http://link.aip.org/link/doi/10.1063/1.2952290

PUBLICATION DATA

ISSN:

1932-1058 (print)  
1932-1058 (online)

Publisher:

$abstract.society.value is a Member of CrossRef American Institute of Physics

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Figures (click on thumbnails to view enlargements)

FIG.1
Configuration drawing of the transparent cell-culturing device. (a) Shows the cross-sectional top view, front view, and side view around the culture region. In the front view the cross section of the gate structure is shown. The arrangement of the upstream/downstream gate and the culture region is in the top view. In the side view the medium flow path and the effusion hole is shown. (b) The whole view of the entire device.

FIG.1 Download High Resolution Image (.zip file) | Export Figure to PowerPoint

FIG.2
Flow simulation of the gated flow chamber at equilibrium. (a) Flow in the chamber with gates and velocity distribution in transversal (A-A’ (b)); longitudinal (B-B’ (c)) directions. The flow rate at the inlet is 10 μL/h. Due to the gated design, the velocity profile inside the culture chamber is uniform over 90% of the culture area. The velocity was calculated at 2 μm above the hole-side and cover-side surface (z = 2 μm).

FIG.2 Download High Resolution Image (.zip file) | Export Figure to PowerPoint

FIG.3
Simulation of the medium replacement. (a) Start of the replacement, t = 0 min. (b) Replacement progression at t = 240 min. (c) Complete replacement at t = 800 min. Medium flow rate is 10 μL/h from left to right. (d) Video showing the simulated medium replacement (enhanced online).

FIG.3 Download High Resolution Image (.zip file) | Export Figure to PowerPoint

FIG.4
Experimental result of the medium replacement. (a) Start of the replacement, t = 0 min. (b) Replacement progression at t = 240 min and (c) t = 270 min. (d) Complete replacement at t = 860 min. The medium flow is 10 μL/h from left to right. (e) Video showing the experimental medium replacement (enhanced online).

FIG.4 Download High Resolution Image (.zip file) | Export Figure to PowerPoint

FIG.5
Simulation of chemical diffusion in a pre-established flow field. The chemical effuses from the central effusion hole at the bottom of the cell-culture chamber at rate of 10 μL/h. (a) Start of the effusion, t = 0 min. (b) Gradient build-up progression at t = 32 min. (c) Steady gradient at t = 128 min. Medium flow is 10 μL/h from left to right. (d) is the transversal gradient generated in (c). Wide segments denote the near-linear region. The “x” denotes the downstream distance from the effusion hole. (e) Video showing the animated simulation of the gradient progression (enhanced online).

FIG.5 Download High Resolution Image (.zip file) | Export Figure to PowerPoint

FIG.6
(a) Simulated flow velocity profile in transversal direction along the A-A’ line in Fig. 5a. The x and y directions are as depicted in Fig. 5a. (b) A plot showing that flow field variation is below 10% at 5 mm downstream away from the effusion hole (x = 5). The flow speed variation is large near the effusion hole and stabilizes gradually as distance from the hole increases.

FIG.6 Download High Resolution Image (.zip file) | Export Figure to PowerPoint

FIG.7
Experimental result of chemical gradient progression. (a) t = 0 min. (b) t = 32 min. (c) t = 128 min. The lateral medium flow is 10 μL/h from left to right. Dye effusion from the inlet is 10 μL/h. (d)–(f) Gradient with a larger medium flow of 30 μL/h, showing a narrower dye distribution. (g)–(i) Gradient with a 50 μL/h lateral medium flow.

FIG.7 Download High Resolution Image (.zip file) | Export Figure to PowerPoint

FIG.8
(a) Temperature distribution in the cell-culture region recorded by an IR thermal imager. Steady medium flow (100 μL/h) is pumped through the culture chamber. Temperature homogeneity is kept to be within ±1 °C in an area of 10 mm×19 mm. (b) Long-term temperature stability of the culture chamber.

FIG.8 Download High Resolution Image (.zip file) | Export Figure to PowerPoint

FIG.9
Image of lung cancer cells, CL1-5, cultured in the microfluidic device after 122 h. The medium flow rate is 10 μL/h. The chamber temperature is kept at 37±1 °C. The inset shows the region where the image was taken.

FIG.9 Download High Resolution Image (.zip file) | Export Figure to PowerPoint

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