[Problem] Provided is a method for detecting of an autofluorescence Liquid Biopsy of Methylated Fragmented DNA (fragmented nucleosome) released into the blood by cell apoptosis as a disease-related substance

[Solution] The inventive method comprises a) a step of capturing the fragmented DNA (fragmented nucleosome) in the analyte as a disease-related substance onto the plasmonic metal meso-crystals; b) a step of irradiating the captured fragmented DNA (fragmented nucleosome) on the plasmonic metal meso-crystal with excitation light to enhance the autofluorescence by the surface plasmon enhancing effect, and acquiring a fluorescent colony image via a filter in a longer wavelength range than the excitation light filter; c) a step of adopting a pixel that exhibits a brightness greater than or equal to a predetermined threshold value of said fluorescent colony image; d) calculating a ratio of a total area value of pixels greater than or equal to a predetermined threshold value of a different two-wavelength region of the adopted measurement region.

[Problem] Provided is a method for detecting of an autofluorescence Liquid Biopsy of Methylated Fragmented DNA (fragmented nucleosome) released into the blood by cell apoptosis as a disease-related substance

[Solution] The inventive method comprises a) a step of capturing the fragmented DNA (fragmented nucleosome) in the analyte as a disease-related substance onto the plasmonic metal meso-crystals; b) a step of irradiating the captured fragmented DNA (fragmented nucleosome) on the plasmonic metal meso-crystal with excitation light to enhance the autofluorescence by the surface plasmon enhancing effect, and acquiring a fluorescent colony image via a filter in a longer wavelength range than the excitation light filter; c) a step of adopting a pixel that exhibits a brightness greater than or equal to a predetermined threshold value of said fluorescent colony image; d) calculating a ratio of a total area value of pixels greater than or equal to a predetermined threshold value of a different two-wavelength region of the adopted measurement region.

An embodiment of a system includes a compartment-generating device, a compartment detector, and electronic computing circuitry. The device is configured to generate compartments of a digital assay, at least one of the compartments having a respective volume that is different from a respective volume of each of at least another one of the compartments. The detector is configured to determine a number of the compartments each having a respective number of a target that is greater than a threshold number of the target. And the electronic circuitry is configured to determine a bulk concentration of the target in a source of the sample in response to the determined number of compartments. Because such a system can be configured to estimate a bulk concentration of a target in a source from a polydisperse digital assay, the system can be portable, and lower-cost and faster, than conventional systems.

An embodiment of a system includes a compartment-generating device, a compartment detector, and electronic computing circuitry. The device is configured to generate compartments of a digital assay, at least one of the compartments having a respective volume that is different from a respective volume of each of at least another one of the compartments. The detector is configured to determine a number of the compartments each having a respective number of a target that is greater than a threshold number of the target. And the electronic circuitry is configured to determine a bulk concentration of the target in a source of the sample in response to the determined number of compartments. Because such a system can be configured to estimate a bulk concentration of a target in a source from a polydisperse digital assay, the system can be portable, and lower-cost and faster, than conventional systems.

In some embodiments, a non-transitory processor-readable medium stores code representing instructions to be executed by a processor. The code includes code to cause the processor to receive a plurality of sets of images associated with a sample treated with fluorescence in situ hybridization (FISH) probes. Each image from that set of images is associated with a different focal length using a fluorescence microscope. Each FISH probe can selectively bind to a unique location on chromosomal DNA in the sample. The code further causes the processor to identify cell nuclei in the images. The code further causes the processor to apply a convolutional neural network (CNN) to each set of images. The CNN is configured to identify a probe indication from a plurality of probe indications for that set of images. The code further causes the processor to identify the sample as containing circulating tumor cells.

The present disclosure generally relates, in certain aspects, to droplet-based microfluidic devices and methods. In certain aspects, target nucleic acids contained within droplets are amplified within droplets in a first step, where multiple primers may be present. However, multiple primers may cause multiple target nucleic acids to be amplified within the droplets, which can make it difficult to identify which nucleic acids were amplified. In a second step, the amplified nucleic acids may be determined. For example, the droplets may be broken and the amplified nucleic acids can be pooled together and sequenced. As an example, new droplets may be formed containing the amplified nucleic acids, and those nucleic acids within the droplets amplified by exposure to certain primers.

The present disclosure generally relates, in certain aspects, to droplet-based microfluidic devices and methods. In certain aspects, target nucleic acids contained within droplets are amplified within droplets in a first step, where multiple primers may be present. However, multiple primers may cause multiple target nucleic acids to be amplified within the droplets, which can make it difficult to identify which nucleic acids were amplified. In a second step, the amplified nucleic acids may be determined. For example, the droplets may be broken and the amplified nucleic acids can be pooled together and sequenced. As an example, new droplets may be formed containing the amplified nucleic acids, and those nucleic acids within the droplets amplified by exposure to certain primers.

The present invention relates to the field of ribonucleotide analysis. More specifically, the present invention provides compositions and methods for detection for nucleic acids using probe pair litigation. In particular embodiments, the compositions and methods of the present invention utilize a probe set comprising (1) a first multi-partite probe comprising a 5′ phosphorylated donor probe and a first bridge probe, wherein the 5′ phosphorylated donor probe specifically hybridizes to a target nucleic acid; and (ii) a second multi-partite probe comprising a 3′ acceptor probe and a second bridge probe, wherein the 3′ acceptor probe specifically hybridizes to the target nucleic acid adjacent to the 5′ donor probe and the second bridge probe is 5′ phosphorylated.

The present invention relates to the field of ribonucleotide analysis. More specifically, the present invention provides compositions and methods for detection for nucleic acids using probe pair litigation. In particular embodiments, the compositions and methods of the present invention utilize a probe set comprising (1) a first multi-partite probe comprising a 5′ phosphorylated donor probe and a first bridge probe, wherein the 5′ phosphorylated donor probe specifically hybridizes to a target nucleic acid; and (ii) a second multi-partite probe comprising a 3′ acceptor probe and a second bridge probe, wherein the 3′ acceptor probe specifically hybridizes to the target nucleic acid adjacent to the 5′ donor probe and the second bridge probe is 5′ phosphorylated.

Provided is a method for detecting and quantifying a nucleic acid in real time and at high speed. The present disclosure provides a real-time high-speed PCR method in which fluorescent signals can be measured from a single-wavelength light source by using a single signal fluorescent material under continuous temperature control. Thus, the PCR method can be performed with a compact lightweight device with a simplified structure.