ac7b03834_si_001.mpg (9.15 MB)
A Multifunctional Reactor with Dry-Stored Reagents for Enzymatic Amplification of Nucleic Acids
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posted on 2017-12-11, 00:00 authored by Jinzhao Song, Changchun Liu, Michael G. Mauk, Jing Peng, Thomas Schoenfeld, Haim H. BauTo
enable inexpensive molecular detection at the point-of-care
and at home with minimal or no instrumentation, it is necessary to
streamline unit operations and store reagents refrigeration-free.
To address this need, a multifunctional enzymatic amplification reactor
that combines solid-phase nucleic acid extraction, concentration,
and purification; refrigeration-free storage of reagents with just-in-time
release; and enzymatic amplification is designed, prototyped, and
tested. A nucleic acid isolation membrane is placed at the reactor’s
inlet, and paraffin-encapsulated reagents are prestored within the
reactor. When a sample mixed with chaotropic agents is filtered through
the nucleic acid isolation membrane, the membrane binds nucleic acids
from the sample. Importantly, the sample volume is decoupled from
the reaction volume, enabling the use of relatively large sample volumes
for high sensitivity. When the amplification reactor’s temperature
increases to its operating level, the paraffin encapsulating the reagents
melts and moves out of the way. The reagents are hydrated, just-in-time,
and the polymerase reaction proceeds. The amplification process can
be monitored, in real-time. We demonstrate our reactors’ ability
to amplify both DNA and RNA targets using polymerase with both reverse-transcriptase
and strand displacement activities to obtain sensitivities on-par
with benchtop equipment and a shelf life exceeding 6 months.
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polymerase reaction proceedssample volumesstrand displacement activitiesparaffin-encapsulated reagentsacid extractionsensitivityparaffin encapsulatingshelf life6 monthssample volumeRNA targetssensitivities on-parreaction volumeamplification processacid isolation membranestore reagents refrigeration-freeunit operationsDNArefrigeration-free storageDry-Stored ReagentsNucleic AcidsMultifunctional Reactorchaotropic agentsbenchtop equipmentEnzymatic Amplificationamplification reactor
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