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Digital LAMP in a sample self-digitization (SD) chip.

ABSTRACT: This paper describes the realization of digital loop-mediated DNA amplification (dLAMP) in a sample self-digitization (SD) chip. Digital DNA amplification has become an attractive technique to quantify absolute concentrations of DNA in a sample. While digital polymerase chain reaction is still the most widespread implementation, its use in resource-limited settings is impeded by the need for thermal cycling and robust temperature control. In such situations, isothermal protocols that can amplify DNA or RNA without thermal cycling are of great interest. Here, we accomplished the successful amplification of single DNA molecules in a stationary droplet array using isothermal digital loop-mediated DNA amplification. Unlike most (if not all) existing methods for sample discretization, our design allows for automated, loss-less digitization of sample volumes on-chip. We demonstrated accurate quantification of relative and absolute DNA concentrations with sample volumes of less than 2 ?l. We assessed the homogeneity of droplet size during sample self-digitization in our device, and verified that the size variation was small enough such that straightforward counting of LAMP-active droplets sufficed for data analysis. We anticipate that the simplicity and robustness of our SD chip make it attractive as an inexpensive and easy-to-operate device for DNA amplification, for example in point-of-care settings.

SUBMITTER: Gansen A

PROVIDER: S-EPMC3383853 | biostudies-literature | 2012 Jun

REPOSITORIES: biostudies-literature

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Digital LAMP in a sample self-digitization (SD) chip.

Gansen Alexander A Herrick Alison M AM Dimov Ivan K IK Lee Luke P LP Chiu Daniel T DT

Lab on a chip 20120307 12

This paper describes the realization of digital loop-mediated DNA amplification (dLAMP) in a sample self-digitization (SD) chip. Digital DNA amplification has become an attractive technique to quantify absolute concentrations of DNA in a sample. While digital polymerase chain reaction is still the most widespread implementation, its use in resource-limited settings is impeded by the need for thermal cycling and robust temperature control. In such situations, isothermal protocols that can amplify ...[more]

PMID: 22399016

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Project description:Digital polymerase chain reaction (dPCR) is a powerful tool for genetic analysis, providing superior sensitivity and accuracy. In many applications that demand minuscule reaction volumes, such as single cell analysis, efficient and reproducible sample handling and digitization is pivotal for accurate absolute quantification of targets, but remains a significant technical challenge. In this paper, we described a robust and flexible microfluidic alternating-pull-push active digitization (?APPAD) strategy that confers close to 100% sample digitization efficiency for microwell-based dPCR. Our strategy employs pneumatic valve control to periodically manipulate air pressure inside the chip to greatly facilitate the vacuum-driven partition of solution into microwells, enabling efficient digitization of a small-volume solution with significantly reduced volume variability. The ?APPAD method was evaluated on both tandem-channel and parallel-channel chips, which achieved a digitization efficiency of 99.5 ± 0.3% and 94.6 ± 0.9% within 10.5 min and 2 min, respectively. To assess the analytical performance of the ?APPAD chip, we calibrated it for absolution dPCR quantitation of ?DNA across a range of concentrations. The results obtained with our chip matched well with the theoretical curve computed from Poisson statistics. Compared to the existing methods for highly efficient sample digitization, not only does our technology greatly reduce the constraints on microwell geometries and channel design, but also benefits from the intrinsic amenability of the pneumatic valve technique with device integration and automation. Thus we envision that the ?APPAD technology will provide a scalable and widely adaptable platform to promote the development of advanced lab-on-a-chip systems integrating microscale sample processing with dPCR for a broad scope of applications, such as single cell analysis of tumor heterogeneity and genetic profiling of circulating exosomes directly in clinical samples.

Project description:BackgroundThe recent proliferation and application of digital technologies in public health has spurred interest in digital public health. However, as yet, there appears to be a lack of conceptual clarity and consensus on its definition.ObjectiveIn this scoping review, we seek to assess formal and informal definitions of digital public health in the literature and to understand how these definitions have been conceptualized in relation to digitization, digitalization, and digital transformation.MethodsWe conducted a scoping literature search in Ovid MEDLINE, Embase, Google Scholar, and 14 government and intergovernmental agency websites encompassing 6 geographic regions. Among a total of 409 full articles identified, we reviewed 11 publications that either formally defined digital public health or informally described the integration of digital technologies into public health in relation to digitization, digitalization, and digital transformation, and we conducted a thematic analysis of the identified definitions.ResultsTwo explicit definitions of digital public health were identified, each with divergent meanings. The first definition suggested digital public health was a reimagination of public health using new ways of working, blending established public health wisdom with new digital concepts and tools. The second definition highlighted digital public health as an asset to achieve existing public health goals. In relation to public health, digitization was used to refer to the technical process of converting analog records to digital data, digitalization referred to the integration of digital technologies into public health operations, and digital transformation was used to describe a cultural shift that pervasively integrates digital technologies and reorganizes services on the basis of the health needs of the public.ConclusionsThe definition of digital public health remains contested in the literature. Public health researchers and practitioners need to clarify these conceptual definitions to harness opportunities to integrate digital technologies into public health in a way that maximizes their potential to improve public health outcomes.International registered report identifier (irrid)RR2-10.2196/preprints.27686.

Dataset Information

Digital LAMP in a sample self-digitization (SD) chip.

Publications

Digital LAMP in a sample self-digitization (SD) chip.

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