This invention relates to methods for molecular detection, particularly to methods utilizing target-specific molecular probes. In exemplary embodiments, target-specific molecular probes include single-stranded deoxyribonucleic acid (DNA) or ribonucleic acid (RNA) aptamers. In general, the molecular probe may bind with relatively high specificity to a given target. In one aspect, a method for molecular detection comprises a molecular probe paired to a reporter molecule wherein the molecular probe impairs the amplification of the reporter molecule in the absence of the target molecule.

This invention relates to methods for molecular detection, particularly to methods utilizing target-specific molecular probes. In exemplary embodiments, target-specific molecular probes include single-stranded deoxyribonucleic acid (DNA) or ribonucleic acid (RNA) aptamers. In general, the molecular probe may bind with relatively high specificity to a given target. In one aspect, a method for molecular detection comprises a molecular probe paired to a reporter molecule wherein the molecular probe impairs the amplification of the reporter molecule in the absence of the target molecule.

Provided herein are genetic circuits and cell state classifiers for detecting the microRNA profile of a cell. The cell state classifiers of the present disclosure utilize phosphorylation state of a transcription factor to control classifier output. Kinases and phosphatase pairs that function in phosphorylating or dephosphorylating the transcription factor are integrated into the circuit, their expression tuned by the presence of microRNAs of interest (e.g., in a cell). The genetic circuits and cell state classifiers may be used in various applications (e.g., therapeutic or diagnostic applications).

Provided herein are genetic circuits and cell state classifiers for detecting the microRNA profile of a cell. The cell state classifiers of the present disclosure utilize phosphorylation state of a transcription factor to control classifier output. Kinases and phosphatase pairs that function in phosphorylating or dephosphorylating the transcription factor are integrated into the circuit, their expression tuned by the presence of microRNAs of interest (e.g., in a cell). The genetic circuits and cell state classifiers may be used in various applications (e.g., therapeutic or diagnostic applications).

Disclosed are means of stimulating systemic immunity and reduction of post-surgery tumor metastasis through the concurrent intralymphatic inhibition of NR2F6 and treatment with cannabidiol. In some embodiments NR2F6 is inhibited by high pressure transient delivery of short interfering RNA into tumor draining lymph nodes concurrent with systemic administration of cannabidiol. This combination may be performed together with means that induce immunogenic tumor cell death. Through the combination of immunogenic cell death and immune stimulation, the invention provides a means of enhancing the abscopal effect and in some embodiments to cause immunological mediated destruction primary and secondary neoplasia.

The present invention provides a method of detecting one or more analytes in a target sample, the method comprising: a. providing a nanoparticle dimer adapted to bind the analyte; b. causing the dimer to pass through a nanopore by voltage-driven translocation; c. observing changes in the translocation current; and d. comparing the translocation current profile of the target sample to the translocation current profile of a control sample; wherein a change in the translocation current profile of the target sample versus the control sample indicates the presence of the analyte in the target sample. Also provided is a method of detecting one or more analytes in a target sample, the method comprising: a. providing a nanoparticle adapted to bind the analyte; b. providing a carrier nucleic acid molecule with at least one single-stranded region; c. contacting the carrier nucleic acid molecule and nanoparticle with the target sample, forming a carrier nucleic acid/analyte/nanoparticle complex; b. causing the carrier nucleic acid/analyte/nanoparticle complex to pass through a biological nanopore by voltage-driven translocation; c. observing changes in the translocation current; and d. comparing the translocation current profile of the target sample to the translocation current profile of a control sample; wherein a change in the translocation current profile of the target sample versus the control sample indicates the presence of the analyte in the target sample.

The present invention provides a method of detecting one or more analytes in a target sample, the method comprising: a. providing a nanoparticle dimer adapted to bind the analyte; b. causing the dimer to pass through a nanopore by voltage-driven translocation; c. observing changes in the translocation current; and d. comparing the translocation current profile of the target sample to the translocation current profile of a control sample; wherein a change in the translocation current profile of the target sample versus the control sample indicates the presence of the analyte in the target sample. Also provided is a method of detecting one or more analytes in a target sample, the method comprising: a. providing a nanoparticle adapted to bind the analyte; b. providing a carrier nucleic acid molecule with at least one single-stranded region; c. contacting the carrier nucleic acid molecule and nanoparticle with the target sample, forming a carrier nucleic acid/analyte/nanoparticle complex; b. causing the carrier nucleic acid/analyte/nanoparticle complex to pass through a biological nanopore by voltage-driven translocation; c. observing changes in the translocation current; and d. comparing the translocation current profile of the target sample to the translocation current profile of a control sample; wherein a change in the translocation current profile of the target sample versus the control sample indicates the presence of the analyte in the target sample.

A liquid chromatography method for simultaneously detecting multiple microRNAs based on a duplex-specific nuclease (DSN) cyclic amplification technology comprises the following steps: designing a fluorophore-modified single-stranded DNA probe according to a target microRNA to be detected and loading the probe onto a surface of a streptavidin-coated magnetic bead (MB) to serve as a detection probe; adding a target microRNA sample to be detected and DSN to the detection probe, fully mixing the same, and incubating the mixture; after the incubation, completely removing the magnetic bead and the unreacted DNA probe to obtain a separated solution; and injecting the separated solution into a high-performance liquid chromatography system for separation and quantification.

This invention relates to a bioassay method for detecting and/or quantitating a short single-stranded nucleic acid analyte employing a binary probe system, where at least one of the two discrete oligonucleotide probe parts of the binary probe has partially double-stranded (self-complementary) stem-loop structure at one terminus and single-stranded overhang sequence region at the other terminus, where the single-stranded terminal regions of both discrete parts of the binary probe hybridize to adjacent complementary regions in the sequence of the nucleic acid analyte molecule, and at least one discrete part of the binary probe comprising a stem-loop structure and single-stranded overhang sequence region hybridizes to terminal region in the sequence of the nucleic acid analyte molecule forming a nick structure. The binary probe system employed in the bioassay method is based on a luminescent reporter technology, either lanthanide chelate complementation or resonance energy transfer with lanthanide label as a donor. Thereby the method allows detection and/or quantitation of the short nucleic acid analyte molecule by time-resolved fluorometry.

This invention relates to a bioassay method for detecting and/or quantitating a short single-stranded nucleic acid analyte employing a binary probe system, where at least one of the two discrete oligonucleotide probe parts of the binary probe has partially double-stranded (self-complementary) stem-loop structure at one terminus and single-stranded overhang sequence region at the other terminus, where the single-stranded terminal regions of both discrete parts of the binary probe hybridize to adjacent complementary regions in the sequence of the nucleic acid analyte molecule, and at least one discrete part of the binary probe comprising a stem-loop structure and single-stranded overhang sequence region hybridizes to terminal region in the sequence of the nucleic acid analyte molecule forming a nick structure. The binary probe system employed in the bioassay method is based on a luminescent reporter technology, either lanthanide chelate complementation or resonance energy transfer with lanthanide label as a donor. Thereby the method allows detection and/or quantitation of the short nucleic acid analyte molecule by time-resolved fluorometry.