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Digital PCR on a SlipChip

Shen, Feng and Du, Wenbin and Kreutz, Jason E. and Fok, Alice and Ismagilov, Rustem F. (2010) Digital PCR on a SlipChip. Lab on a Chip, 10 (20). pp. 2666-2672. ISSN 1473-0197. PMCID PMC2948063. https://resolver.caltech.edu/CaltechAUTHORS:20130821-160730931

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Abstract

This paper describes a SlipChip to perform digital PCR in a very simple and inexpensive format. The fluidic path for introducing the sample combined with the PCR mixture was formed using elongated wells in the two plates of the SlipChip designed to overlap during sample loading. This fluidic path was broken up by simple slipping of the two plates that removed the overlap among wells and brought each well in contact with a reservoir preloaded with oil to generate 1280 reaction compartments (2.6 nL each) simultaneously. After thermal cycling, end-point fluorescence intensity was used to detect the presence of nucleic acid. Digital PCR on the SlipChip was tested quantitatively by using Staphylococcus aureus genomic DNA. As the concentration of the template DNA in the reaction mixture was diluted, the fraction of positive wells decreased as expected from the statistical analysis. No cross-contamination was observed during the experiments. At the extremes of the dynamic range of digital PCR the standard confidence interval determined using a normal approximation of the binomial distribution is not satisfactory. Therefore, statistical analysis based on the score method was used to establish these confidence intervals. The SlipChip provides a simple strategy to count nucleic acids by using PCR. It may find applications in research applications such as single cell analysis, prenatal diagnostics, and point-of-care diagnostics. SlipChip would become valuable for diagnostics, including applications in resource-limited areas after integration with isothermal nucleic acid amplification technologies and visual readout.


Item Type:Article
Related URLs:
URLURL TypeDescription
https://doi.org/10.1039/c004521gDOIArticle
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2948063PubMed CentralArticle
ORCID:
AuthorORCID
Shen, Feng0000-0002-4709-330X
Du, Wenbin0000-0002-7401-1410
Ismagilov, Rustem F.0000-0002-3680-4399
Additional Information:© Royal Society of Chemistry 2010. Received 22nd March 2010, Accepted 9th June 2010. First published on the web 1st July 2010. This work was supported by ONR grant No. N00014-08-1-0936, the Camille Dreyfus Teacher-Scholar Awards program, the Chicago Center for Systems Biology (funded by the National Institute of General Medical Sciences at the NIH), and by the NIH Director's Pioneer Award program, part of the NIH Roadmap for Medical Research (1 DP1 OD003584). Part of this work was performed at the Materials Research Science and Engineering Centers microfluidics facility (funded by the National Science Foundation). We thank Elena M. Lucchetta for additional experimental validation and Elena K. Davydova for helpful suggestions in experimental design. We thank Heidi Park for contributions to writing and editing this manuscript.
Funders:
Funding AgencyGrant Number
Office of Naval Research (ONR)N00014-08-1-0936
Camille and Henry Dreyfus FoundationUNSPECIFIED
National Institute of General Medical SciencesUNSPECIFIED
NIH1DP1 OD003584
NSFUNSPECIFIED
Issue or Number:20
PubMed Central ID:PMC2948063
Record Number:CaltechAUTHORS:20130821-160730931
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20130821-160730931
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:40865
Collection:CaltechAUTHORS
Deposited By: Whitney Barlow
Deposited On:26 Aug 2013 22:08
Last Modified:04 Mar 2020 23:47

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