The NTD Nanoscope: potential applications and implementations

Abstract

BackgroundNanopore transduction detection (NTD) offers prospects for a number of highly sensitive and discriminative applications, including: (i) single nucleotide polymorphism (SNP) detection; (ii) targeted DNA re-sequencing; (iii) protein isoform assaying; and (iv) biosensing via antibody or aptamer coupled molecules. Nanopore event transduction involves single-molecule biophysics, engineered information flows, and nanopore cheminformatics. The NTD Nanoscope has seen limited use in the scientific community, however, due to lack of information about potential applications, and lack of availability for the device itself. Meta Logos Inc. is developing both pre-packaged device platforms and component-level (unassembled) kit platforms (the latter described here). In both cases a lipid bi-layer workstation is first established, then augmentations and operational protocols are provided to have a nanopore transduction detector. In this paper we provide an overview of the NTD Nanoscope applications and implementations. The NTD Nanoscope Kit, in particular, is a component-level reproduction of the standard NTD device used in previous research papers.ResultsThe NTD Nanoscope method is shown to functionalize a single nanopore with a channel current modulator that is designed to transduce events, such as binding to a specific target. To expedite set-up in new lab settings, the calibration and troubleshooting for the NTD Nanoscope kit components and signal processing software, the NTD Nanoscope Kit, is designed to include a set of test buffers and control molecules based on experiments described in previous NTD papers (the model systems briefly described in what follows). The description of the Server-interfacing for advanced signal processing support is also briefly mentioned.ConclusionsSNP assaying, SNP discovery, DNA sequencing and RNA-seq methods are typically limited by the accuracy of the error rate of the enzymes involved, such as methods involving the polymerase chain reaction (PCR) enzyme. The NTD Nanoscope offers a means to obtain higher accuracy as it is a single-molecule method that does not inherently involve use of enzymes, using a functionalized nanopore instead.

Channel current cheminformatics and stochastic carrier-wave signal processing

“…The stochastic sequential analysis methods, described in what follows, provide a robust and efficient means to enhance a device or process, as well as possibly enabling new capabilities for the device or process (via transduction coupling, for example, as with the nanopore transduction detector (NTD) platform [22]). The SSA Protocol could potentially work with existing device or process information flows, or could work with additional information induced via modulation or via transduction couplings (comprising carrier references [1], among other things).…”

“…In the Nanopore Transduction Detector (NTD) Nanoscope experiments [22], the molecular dynamics of a (single) captured non-translocating transducer molecule provide a unique stochastic reference signal with stable statistics on the observed, single-molecule blockaded channel current, somewhat analogous to a carrier signal in standard electrical engineering signal analysis. Discernible changes in blockade statistics, coupled to SSA signal processing protocols, enable the means for a highly detailed characterization of the interactions of the transducer molecule with binding targets (cognates) in the surrounding (extra-channel) environment.…”

“…The transducer molecule is engineered to generate distinct channel blockade signals depending on its interaction with target molecules [22]. Statistical models are trained for each binding mode, bound and unbound, for example, by exposing the transducer molecule to zero or high (excess) concentrations of the target molecule.…”

“…In addition, the design of the machine learning based algorithms allow for scaling to large datasets, via real-time distributed processing, and are adaptable to analysis on any stochastic sequential dataset. The machine learning software has also been integrated into the NTD Nanoscope [22] for "realtime" pattern-recognition informed (PRI) feedback [45]. The methods used to implement the PRI feedback include distributed HMM and SVM implementations, which enable the processing speedup that is needed.…”

“…A description of the SSA Protocol and SCW communications will be given, where stochastic phase modulation (SPM), a simple form of SCW communication, has been used in engineered nanopore transduction detector experiments [1,22]. A description for a pragmatic distributed HMM generalization and implementation is also given.…”