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Low-voltage paper isotachophoresis device for DNA focusing.

ABSTRACT: We present a new paper-based isotachophoresis (ITP) device design for focusing DNA samples having lengths ranging from 23 to at least 1517 bp. DNA is concentrated by more than two orders of magnitude within 4 min. The key component of this device is a 2 mm-long, 2 mm-wide circular paper channel formed by concertina folding a paper strip and aligning the circular paper zones on each layer. Due to the short channel length, a high electric field of ~16 kV m(-1) is easily generated in the paper channel using two 9 V batteries. The multilayer architecture also enables convenient reclamation and analysis of the sample after ITP focusing by simply opening the origami paper and cutting out the desired layers. We profiled the electric field in the origami paper channel during ITP experiments using a nonfocusing fluorescent tracer. The result showed that focusing relied on formation and subsequent movement of a sharp electric field boundary between the leading and trailing electrolyte.

SUBMITTER: Li X

PROVIDER: S-EPMC4589534 | biostudies-literature | 2015 Oct

REPOSITORIES: biostudies-literature

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Low-voltage paper isotachophoresis device for DNA focusing.

Li Xiang X Luo Long L Crooks Richard M RM

Lab on a chip 20150904 20

We present a new paper-based isotachophoresis (ITP) device design for focusing DNA samples having lengths ranging from 23 to at least 1517 bp. DNA is concentrated by more than two orders of magnitude within 4 min. The key component of this device is a 2 mm-long, 2 mm-wide circular paper channel formed by concertina folding a paper strip and aligning the circular paper zones on each layer. Due to the short channel length, a high electric field of ~16 kV m(-1) is easily generated in the paper chan ...[more]

PMID: 26338530

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Project description:Nucleic acid amplification tests (NAATs) are a crucial diagnostic and monitoring tool for infectious diseases. A key procedural step for NAATs is sample preparation: separating and purifying target nucleic acids from crude biological samples prior to nucleic acid amplification and detection. Traditionally, sample preparation has been performed with liquid- or solid-phase extraction, both of which require multiple trained user steps and significant laboratory equipment. The challenges associated with sample preparation have limited the dissemination of NAAT point-of-care diagnostics in low resource environments, including low- and middle-income countries. We report on a paper-based device for purification of nucleic acids from whole blood using isotachophoresis (ITP) for point-of-care NAATs. We show successful extraction and purification of target nucleic acids from large volumes (33 µL) of whole human blood samples with no moving parts and few user steps. Our device utilizes paper-based buffer reservoirs to fully contain the liquid ITP buffers and does not require complex filling procedures, instead relying on the natural wicking of integrated paper membranes. We perform on-device blood fractionation via filtration to remove leukocytes and erythrocytes from our sample, followed by integrated on-paper proteolytic digestion of endogenous plasma proteins to allow for successful isotachophoretic extraction. Paper-based isotachophoresis purifies and concentrates target nucleic acids that are added directly to recombinase polymerase amplification (RPA) reactions. We show consistent amplification of input copy concentrations of as low as 3 × 103 copies nucleic acid per mL input blood with extraction and purification taking only 30 min. By employing a paper architecture, we are able to incorporate these processes in a single, robust, low-cost design, enabling the direct processing of large volumes of blood, with the only intermediate user steps being the removal and addition of tape. Our device represents a step towards a simple, fully integrated sample preparation system for nucleic acid amplification tests at the point-of-care.

Dataset Information

Low-voltage paper isotachophoresis device for DNA focusing.

Publications

Low-voltage paper isotachophoresis device for DNA focusing.

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