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Increased density and coverage uniformity of viruses on a sensor surface by using U-type, T-type, and W-type microfluidic devices
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10.1063/1.4722294
/content/aip/journal/bmf/6/2/10.1063/1.4722294
http://aip.metastore.ingenta.com/content/aip/journal/bmf/6/2/10.1063/1.4722294

Figures

Image of FIG. 1.
FIG. 1.

Schematic diagram of the folded microfluidic devices There are inlet channels from the right and outlet channels to the left for three folded microfluidic devices. The reaction are of three devices are all 3 mm × 3 mm × 100 μm in the central part of channels. (a) T- type device. No partitions were set up in the T-type microchannel. (b) U- type device. A partition (200 μm × 1.5 mm × 100 μm) was set up in the U-type microchannel dividing the microchannel into two sections of 1.4 mm × 100 μm. (c) W- type device. Three partitions (200 μm × 2.4 mm × 100 μm) were set up in the W-type microchannel dividing the channel into four sections of 600 μm × 100 μm.

Image of FIG. 2.
FIG. 2.

Experimental streamlines in (a) T-type device. In the fluorescent flow line verification of the T-type microchannel, the closer to the inlet section of the central flow axis, the denser the flow line. The flow in the area below the flow lines was thinner and slower. We discovered a problem with bubbles. (b) U-type device. In the fluorescent flow line verification of the U-type microchannel, most of the fluorescent particles were obstructed by the partition; (c) W-type device. In the fluorescent flow line verification of the W-type microchannel, the structure of the microchannel produced characteristics of a high-intensity flow line and motion path. (Image from videos.) (enhanced online)., (c), (d) [URL: http://dx.doi.org/10.1063/1.4722294.1] [URL: http://dx.doi.org/10.1063/1.4722294.2] [URL: http://dx.doi.org/10.1063/1.4722294.3]10.1063/1.4722294.110.1063/1.4722294.210.1063/1.4722294.3

Image of FIG. 3.
FIG. 3.

The carbodiimide group (N=C=N) in NCD-4 can be captured by carboxyl group in 11-MUA. The attachment process is illustrated in Figs. 3(I), ???, ???, and ???–3(IV). (I) A glass substrate was cleaned by ethanol and acetone. (II) Gold was evaporated on a glass substrate. (III) The dipping method or microchannel circulation system was applied to the substrate at room temperature. A layer of 11-MUA self-assembled monolayers was deposited on the substrate over 10 h with a Moore volume concentration of 18.32 mM. (IV) In a low-temperature environment (−20 °C), NCD-4 solution was soaked onto the substrate for 20 h with a Moore volume concentration of 854.99 nM.

Image of FIG. 4.
FIG. 4.

The succinimidyl-4 -(N-maleimidomethyl)cyclohexane-1-carboxylate could conjugate with the linker which has the amino group derived from lysine. After the reaction, the maleimide-activated linker can link to the thiol group of TYMV particle. The viral attachment process is illustrated in Figs. 4(I), ???, ???, and ???. (I) A layer of Au film was deposited on the substrate by sputtering. (II) 20 mM 11-MUA solution was injected on the sensor surface by the microfluidics system. This linked the 11-MUA with gold films. (III) EDC and NHS solutions (molar ratio 2:1) were also injected on the sensor surface by the microfluidics system. (IV) TYMV particles with fluorescent particles (NCD-4) in the buffer were introduced over the sensor surface.

Image of FIG. 5.
FIG. 5.

Confocal fluorescence 2D (left) and 3D (right) micrographs of MUA/NCD4 (a) soaking method. The average fluorescence was 996.34 A.U. and the average fluorescence coverage is 33.37%. Without a considerable concentration gradient in the liquid or random movement patterns, molecular diffusion and Brownian motion lead to intermolecular aggregation. (b) T-type devices. The average fluorescence intensity was 1558.08 A.U., and the average fluorescence coverage was 43.73%.The liquid was nearly stationary in the two lower corners of the T device, and the attached results indicated sporadic reaction. (c) U-type devices. Compared to T-device, the attached results showed more uniform fluorescence distribution in the U-device. The average fluorescence intensity was 2171.43 A.U, and the average fluorescence coverage was 62.34%. (d) W-tape devices. The average fluorescence intensity was 2658.33 A.U. and average fluorescence coverage was 85.13%. Compared to the T-type and U-type microchannels using the traditional immersion method, these results exhibit a substantial improvement in 11-MUA and NCD-4 attachment efficiency.

Image of FIG. 6.
FIG. 6.

Fluorescent intensity and flow rate (a) T-type devices. At a fluorescence strength of x = 1.2 mm, the intensity indicated that fluorescence intensity and simulated flow were mostly positively correlated and may be related to the number of particles in the unit cross-sectional area. (b) U-type devices. At a fluorescence strength of x = 1.2 mm, the intensity is mostly positively correlated to the simulated flow from y = 0 to y = 3 mm, as shown in Fig. 6(b). The negatively correlated zone is near 1500 μm because this zone was positioned at a point where the fluid turned. (c) W-type devices. At a fluorescence strength of x = 0.6 m, fluorescence intensity and simulated flow are positively correlated at a flow velocity below 0.0108 m/s.

Image of FIG. 7.
FIG. 7.

The overall average coverage rates at six distinct sections were subsequently used to analyze the attachment efficiency, and the average fluorescence coverage of the area was used to calculate attachment uniformity. The fluorescence image was processed in gray scale by using the 256 color scale method to calculate the average fluorescence coverage of the six sections in the figure. The calculations excluded the partition area. The average fluorescence coverage of the six areas was used to determine the uniformity of coverage, as shown. (a) In the traditional soaking method, the regional fluorescence coverage was approximately 20% to 40%. (b) For T-type microchannels, the regional fluorescence coverage ranged from 10% to 80% because the motion pattern of the microflow caused this structure to exhibit a coverage gap from 10% to 80%. (c) The U-type microchannels exhibited regional fluorescence coverage ranging from 50% to 90%, with the addition of partitions changing the original flow pattern. (d) The W-type microchannels exhibited regional fluorescence coverage of 80% to 90%. (e) Average fluorescent coverage of MUA at six sections for soaking device, T-device, U-device, and W-device are shown.

Image of FIG. 8.
FIG. 8.

Confocal fluorescence 2D (left) and 3D (right) micrographs of TYMV/A594 fluorescent particles. (a) soaking device. Few viruses were attached to the reaction zone through soaking methods. Average fluorescence intensity was 342.46 A.U., and average fluorescence coverage was 4.23%. (b) W-type device. Compared to traditional immersion methods, the experimental results in W-type device indicate that viruses are attached to the reaction zone in a uniform distribution. Average fluorescence intensity was 1229 A.U. and average fluorescence coverage was 85.13% in the W device.

Image of FIG. 9.
FIG. 9.

Fluorescent intensity of TYMV and flow rate in W-type device. Fluorescence intensity and simulated flow were mostly positively correlated in the experiment of TYMV attachment.

Image of FIG. 10.
FIG. 10.

The overall average coverage rates at six distinct sections were subsequently used to analyze viral attachment efficiency. The fluorescence image was processed in gray-scale using the 256 color scale method. The calculations excluded the partition area. (a) In soaking device, the regional fluorescence coverage was approximately 0% to 10%, and the attachment efficiency was the lowest. (b) The W device exhibited regional fluorescence coverage of 70% to 90%. (c) The average fluorescence coverage of the six areas is shown.

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/content/aip/journal/bmf/6/2/10.1063/1.4722294
2012-05-24
2014-04-19
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752b84549af89a08dbdd7fdb8b9568b5 journal.articlezxybnytfddd
Scitation: Increased density and coverage uniformity of viruses on a sensor surface by using U-type, T-type, and W-type microfluidic devices
http://aip.metastore.ingenta.com/content/aip/journal/bmf/6/2/10.1063/1.4722294
10.1063/1.4722294
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