Research ArticleMICROTECHNOLOGY

Self-powered integrated microfluidic point-of-care low-cost enabling (SIMPLE) chip

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Science Advances  22 Mar 2017:
Vol. 3, no. 3, e1501645
DOI: 10.1126/sciadv.1501645
  • Fig. 1 The SIMPLE chip for low-cost, quantitative, and portable nucleic acid testing.

    (A) The simple operation protocol requires minimal handling and no external pumps or power sources. Users may simply drop blood/amplification reagent mix into the inlet and then the chip performs automatic sample preparation. (B) The chip is then incubated on a reusable heat pack, and end-point isothermal digital amplification of nucleic acid is done (RPA). Scale bar, 2 mm. (C) Dye-loaded chip for visualization of microchannels. Red shows fluidic channels. Blue shows the main vacuum battery system. Green shows the auxiliary vacuum battery system. (D) Digital microfluidic patterning enables reagent patterning with common laboratory equipment. Left: After chip bonding. Right: Amplification initiator concentrated asymmetrically using a microapex stencil (see fig. S3 for details). Scale bars, 100 μm (black) and 1 mm (yellow). (E) Side view of the digital plasma separation design, which removes blood cells via sedimentation and skims plasma into dead-end wells for digital amplification. (F) The vacuum battery system frees the chip from external pumps or power sources for pumping. Vacuum is prestored in the large “battery” voids. Fluid is pumped by slowly releasing the prestored vacuum potential via air diffusion through lung-like structures.

  • Fig. 2 Prepatterning of the amplification initiator on the chip.

    (A) The formation of concentrated asymmetric apex reagent thin film of MgOAc creates isolated spots and smaller footprints. Reagents concentrate asymmetrically toward the tip via capillary tension. Small reagent footprints prevent bonding problems and false positives. Scale bar, 200 μm. (B) Prepatterned MgOAc with fluorescein for visualization. Scale bar, 2 mm. There are four steps for patterning: (1) Reagents can be digitized into discrete samples by degas pumping using a patterning stencil. Food dye was used for visualization. Degas pumping works by slowly sucking the liquid when trapped air diffuses into prevacuumed air-permeable silicone (PDMS) material. A trailing air-gap digitizes the reagents into discrete patterns. (2) Asymmetric apex concentration (mean ± SD, n = 16). A.U., arbitrary units. (3) MgOAc adhered to the blank PDMS after top-patterning stencil is peeled off. Scale bar, 100 μm (black). (4) The patterned PDMS is flipped, aligned, and bonded on top of the layer containing microfluidic patterns for the SIMPLE chip. The microwells were reconstituted with water to verify uniformity. Scale bars, 250 μm (black) and 1 mm (yellow).

  • Fig. 3 One-step autonomous sample preparation with the digital plasma separation design.

    (A) This design enables simultaneous plasma separation and sample compartmentalization for digital isothermal amplification. (B) Blood cells drop below the microcliff gap because of sedimentation, and plasma near the top skims into the wells. Black dashed arrows depict air diffusion across the permeable silicone into the vacuum battery, sucking plasma into the microwells. (C) Automatic compartmentalization occurs when the trailing air gap separates the 224 microwells for digital amplification. Scale bar, 2 mm. (D) Fluorescence images of plasma separation of human blood mixed with DNA. DNA fluorescence in the main channels is obstructed by the opacity of blood cells. Scale bar, 500 μm. (E) Smaller microcliff gaps and lower flow speed can remove >95% of blood cells in the microwells. Dashed lines indicate simulation results. Solid dots indicate experimental results (mean ± SD, Pearson correlation = 0.99, n = 6). (F) No hemolysis was observed using our design. Ultrasound lysed blood, centrifuged plasma, and blood were loaded into separate chips, and absorbance in the microwells was recorded.

  • Fig. 4 Vacuum battery on the chip.

    (A) Image of the two on-chip batteries. Channels are filled with dye for visualization only. (B) The basic unit of the vacuum battery system pumps fluid by slowly releasing stored vacuum potential via air diffusion over the vacuum lung structures. Black arrows depict trapped air diffusing across the vacuum lungs, which, in turn, suck fluid in. (C) Equipment-free loading and automatic sample compartmentalization in 10 min (dye). Digital amplification can be performed as soon as compartmentalization is finished. (D) Flow tuning by varying the main battery volume (auxiliary battery constant). Dashed lines indicate simulation results, solid lines indicate fitted result, and solid dots indicate experimental averages [mean ± SD; adjusted R2 = 0.99; P < 0.01, analysis of variance (ANOVA); n = 3]. (E) After opening the seal, the effect of the time gap before loading samples on the time it takes for the liquid to reach the end of the device was tested. Right photo shows end-loaded chip. (F) This chip has a long window of operation. It pumps for at least 2.5 hours after opening the vacuum pack (40-min gap time + 110-min loading time) and has higher reliability compared to conventional degas methods [Pearson’s R = 0.60 (black line) and 0.95 (red line); n = 3].

  • Fig. 5 Quantitative digital amplification of nucleic acid.

    (A) Left panel shows the concept of digital amplification. Wells that have at least one or more target templates are amplified, whereas others remain unamplified. One can determine the original template concentration by counting the number of amplified wells. Isothermal nucleic acid amplification was done with RPA. Isothermal heating was done via reusable sodium acetate heat packs. Right panel shows end-point fluorescence images of reactions with different starting concentrations of methicillin-resistant Staphylococcus aureus (MRSA) DNA spiked in human whole blood. Scale bar, 5 mm. (B) The average intensity of positive spots increases to a detectable level within 18 min (signal difference >3× SD; mean ± SD; power = 0.99; P < 0.05, two-tailed t test; n = 5). In this test, HIV-1 RNA was spiked in human whole blood. (C) Quantification range of the SIMPLE chip. MRSA DNA was spiked into human whole blood for these tests (mean ± SD, solid line from fitting, adjusted R2 = 0.99, n = 3).

Supplementary Materials

  • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/3/3/e1501645/DC1

    fig. S1. Exploded view of the simple construction.

    fig. S2. Vacuum charging and long-term storage.

    fig. S3. Detailed steps of micropatterning.

    fig. S4. Digital plasma separation mechanism overview.

    fig. S5. Failure of blood separation without the microcliff.

    fig. S6. Optical signal obstructed when blood cells are not removed.

    fig. S7. Microcliff gap effect on spatial robustness.

    fig. S8. Reliable compartmentalization with smaller microcliff gap designs.

    fig. S9. Selective particle separation according to size.

    fig. S10. Total plasma volume separated versus time.

    fig. S11. Microwell filling speed versus vacuum strength.

    fig. S12. Vacuum battery system versus conventional degas pumping.

    fig. S13. Vacuum lungs enable flow tuning.

    fig. S14. Compartmentalization of all 224 microwells can be done in 12 min.

    fig. S15. Consistent loading with the vacuum battery system.

    fig. S16. RPA is more robust against plasma samples than LAMP and PCR.

    fig. S17. Isothermal heating using reusable instant heat packs.

    fig. S18. On-chip digital quantitative detection of MRSA DNA spiked in water.

    note S1. Simulation of particle trajectories.

    note S2. Diffusion through vacuum battery.

    table S1. Primer, probe, and target sequences.

    movie S1. User protocol.

    movie S2. Cross section.

    movie S3. Digital plasma separation.

    movie S4. Vacuum battery system.

    References (5356)

  • Supplementary Materials

    This PDF file includes:

    • fig. S1. Exploded view of the simple construction.
    • fig. S2. Vacuum charging and long-term storage.
    • fig. S3. Detailed steps of micropatterning.
    • fig. S4. Digital plasma separation mechanism overview.
    • fig. S5. Failure of blood separation without the microcliff.
    • fig. S6. Optical signal obstructed when blood cells are not removed.
    • fig. S7. Microcliff gap effect on spatial robustness.
    • fig. S8. Reliable compartmentalization with smaller microcliff gap designs.
    • fig. S9. Selective particle separation according to size.
    • fig. S10. Total plasma volume separated versus time.
    • fig. S11. Microwell filling speed versus vacuum strength.
    • fig. S12. Vacuum battery system versus conventional degas pumping.
    • fig. S13. Vacuum lungs enable flow tuning.
    • fig. S14. Compartmentalization of all 224 microwells can be done in 12 min.
    • fig. S15. Consistent loading with the vacuum battery system.
    • fig. S16. RPA is more robust against plasma samples than LAMP and PCR.
    • fig. S17. Isothermal heating using reusable instant heat packs.
    • fig. S18. On-chip digital quantitative detection of MRSA DNA spiked in water.
    • note S1. Simulation of particle trajectories.
    • note S2. Diffusion through vacuum battery.
    • table S1. Primer, probe, and target sequences.
    • References (53–56)

    Download PDF

    Other Supplementary Material for this manuscript includes the following:

    • movie S1(.mov format). User protocol.
    • movie S2 (.mov format). Cross section.
    • movie S3 (.mov format). Digital plasma separation.
    • movie S4 (.mov format). Vacuum battery system.

    Files in this Data Supplement:

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