Advertisement

SlipChip Device for Digital Nucleic Acid Amplification

  • Feng ShenEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1547)

Abstract

Digital nucleic acid amplification (Digital NAA) quantifies nucleic acid by compartmentalizing a sample of DNA or RNA into a large number of discrete partitions and performing parallel nucleic acid amplification, such as polymerase chain reaction (PCR) or isothermal amplification reactions. With the counts of positive wells, total number of wells, and volumes of wells, the concentration of the target nucleic acid in the sample can be quantified. Digital NAA is considered increasingly powerful for ultra-sensitive detection and accurate quantification of nucleic acid for biological research and potentially medical diagnostics. Here, we describe glass SlipChip devices to perform digital NAA without cumbersome manual manipulation or complex fluidic control systems.

Key words

Digital PCR Quantification Nucleic acid amplification Microfluidics Lab on a chip Ultra-sensitive detection Single molecule 

References

  1. 1.
    Baker M (2012) Digital PCR hits its stride. Nat Methods 9(6):541–544CrossRefGoogle Scholar
  2. 2.
    Huggett JF et al (2013) The digital MIQE guidelines: minimum information for publication of quantitative digital PCR experiments. Clin Chem 59(6):892–902CrossRefGoogle Scholar
  3. 3.
    Witters D et al (2014) Digital biology and chemistry. Lab Chip 14(17):3225–3232CrossRefGoogle Scholar
  4. 4.
    Hindson BJ et al (2011) High-throughput droplet digital PCR system for absolute quantitation of DNA copy number. Anal Chem 83(22):8604–8610CrossRefGoogle Scholar
  5. 5.
    Ottesen EA, Hong JW, Quake SR, Leadbetter JR (2006) Microfluidic digital PCR enables multigene analysis of individual environmental bacteria. Science 314(5804):1464–1467CrossRefGoogle Scholar
  6. 6.
    Shen F, Du W, Kreutz JE, Fok A, Ismagilov RF (2010) Digital PCR on a SlipChip. Lab Chip 10(20):2666–2672CrossRefGoogle Scholar
  7. 7.
    Shen F et al (2011) Multiplexed quantification of nucleic acids with large dynamic range using multivolume digital RT-PCR on a rotational SlipChip tested with HIV and hepatitis C viral load. J Am Chem Soc 133(44):17705–17712CrossRefGoogle Scholar
  8. 8.
    Shen F et al (2011) Digital isothermal quantification of nucleic acids via simultaneous chemical initiation of recombinase polymerase amplification reactions on SlipChip. Anal Chem 83(9):3533–3540CrossRefGoogle Scholar
  9. 9.
    Sun B et al (2013) Mechanistic evaluation of the pros and cons of digital RT-LAMP for HIV-1 viral load quantification on a microfluidic device and improved efficiency via a two-step digital protocol. Anal Chem 85(3):1540–1546CrossRefGoogle Scholar
  10. 10.
    Du W, Li L, Nichols KP, Ismagilov RF (2009) SlipChip. Lab Chip 9(16):2286–2292CrossRefGoogle Scholar
  11. 11.
    Shen F et al (2010) Nanoliter multiplex pcr arrays on a slipchip. Anal Chem 82(11):4606–4612CrossRefGoogle Scholar
  12. 12.
    Sun B et al (2014) Measuring fate and rate of single-molecule competition of amplification and restriction digestion, and its use for rapid genotyping tested with hepatitis C viral RNA. Angew Chemie Int Ed 53(31):8088–8092CrossRefGoogle Scholar
  13. 13.
    Li L, Du W, Ismagilov R (2010) Multiparameter screening on slipchip used for nanoliter protein crystallization combining free interface diffusion and microbatch methods. J Am Chem Soc 132(1):112–119CrossRefGoogle Scholar
  14. 14.
    Liu W, Chen D, Du W, Nichols KP, Ismagilov RF (2010) SlipChip for immunoassays in nanoliter volumes. Anal Chem 82(8):3276–3282CrossRefGoogle Scholar
  15. 15.
    Ge S, Liu W, Schlappi T, Ismagilov RF (2014) Digital, ultrasensitive, end-point protein measurements with large dynamic range via brownian trapping with drift. J Am Chem Soc 136(42):14662–14665CrossRefGoogle Scholar
  16. 16.
    Begolo S, Shen F, Ismagilov RF (2013) A microfluidic device for dry sample preservation in remote settings. Lab Chip 13(22):4331–4342CrossRefGoogle Scholar
  17. 17.
    Ma L et al (2014) Gene-targeted microfluidic cultivation validated by isolation of a gut bacterium listed in human microbiome project’s most wanted taxa. Proc Natl Acad Sci U S A 111(27):9768–9773CrossRefGoogle Scholar
  18. 18.
    Zhao Y, Pereira F, deMello AJ, Morgan H, Niu X (2014) Droplet-based in situ compartmentalization of chemically separated components after isoelectric focusing in a Slipchip. Lab Chip 14(3):555–561CrossRefGoogle Scholar
  19. 19.
    Li L, Karymov MA, Nichols KP, Ismagilov RF (2010) Dead-end filling of slipchip evaluated theoretically and experimentally as a function of the surface chemistry and the gap size between the plates for lubricated and dry slipchips. Langmuir 26(14):12465–12471CrossRefGoogle Scholar
  20. 20.
    Begolo S, Zhukov DV, Selck DA, Li L, Ismagilov RF (2014) The pumping lid: investigating multi-material 3D printing for equipment-free, programmable generation of positive and negative pressures for microfluidic applications. Lab Chip 14(24):4616–4628CrossRefGoogle Scholar
  21. 21.
    Selck DA, Karymov MA, Sun B, Ismagilov RF (2013) Increased robustness of single-molecule counting with microfluidics, digital isothermal amplification, and a mobile phone versus real-time kinetic measurements. Anal Chem 85(22):11129–11136CrossRefGoogle Scholar
  22. 22.
    Kreutz JE et al (2011) Theoretical design and analysis of multivolume digital assays with wide dynamic range validated experimentally with microfluidic digital PCR. Anal Chem 83(21):8158–8168CrossRefGoogle Scholar
  23. 23.
    Mazutis L et al (2009) Droplet-based microfluidic systems for high-throughput single DNA molecule isothermal amplification and analysis. Anal Chem 81(12):4813–4821CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media LLC 2017

Authors and Affiliations

  1. 1.SlipChip CorporationMenlo ParkUSA

Personalised recommendations