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Encapsulated sensors and sensing systems for bioassays and diagnostics and methods for making and using them

A sensing system and sensor technology, applied in the field of biological analysis, can solve the problem of missing key dynamic information of single cells

Active Publication Date: 2016-07-13
RGT UNIV OF CALIFORNIA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In contrast, existing techniques (e.g. ELISA) typically measure large volumes of secreted proteins and thus miss critical dynamic information at the single-cell level

Method used

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  • Encapsulated sensors and sensing systems for bioassays and diagnostics and methods for making and using them
  • Encapsulated sensors and sensing systems for bioassays and diagnostics and methods for making and using them
  • Encapsulated sensors and sensing systems for bioassays and diagnostics and methods for making and using them

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0288] Example 1: Detection of bacteria in biological samples using microencapsulated sensors

[0289] Real-time fluorescent DNAzyme sensor : In one embodiment, use includes about 10 4 DNA libraries of random sequences (eg, chemically synthesized) are used to select and / or isolate DNAzyme sensors. The library can consist, for example, of approximately 40 nucleotides of variable sequences linked to a fluorescent DNA-RNA chimeric substrate (see Figure 10 a). 7 The substrate may contain a single ribonucleoside (riboadenosine) as a cleavage site flanked on each side by a fluorophore and a quencher, respectively. The rationale is that specific DNA sequences in the library exist only in the presence of target bacterial lysates and are cleaved at nucleotide junctions, thereby generating fluorescent signals.

[0290] In vitro selection can be achieved by incubating the initial library with target bacterial lysates in HEPES buffer for approximately 10 minutes. The cleaved mole...

Embodiment 2

[0325] Example 2: Detection of Bacterial Infection Using 3D Particle Counter-Integrated System (i.e. IC3D)

[0326] Exemplary methods of the present invention for detection of bloodstream infection (BSI), and rapid detection, identification, and treatment of bacteria in the early stages of infection are described below.

[0327] It has been demonstrated that the integrated droplet system and 3D particle counter system of the present invention allows selective detection of bacteria in untreated or minimally processed buffer and blood samples with single cell sensitivity within minutes to hours. In an alternative embodiment, the present system integrates DNAzyme sensor technology, droplet microfluidics, and a high-throughput 3D particle counting system (i.e., Integrated Integrated Droplet Digital Detection (IC3D)) ( figure 1 and figure 2 c). This exemplary combination allows the selective detection of single cells in milliliter volumes of blood within minutes.

[0328] In...

Embodiment 3

[0348] Example 3: Detection of Antimicrobial Resistance by IC3D Using Fluorescent Substrates

[0349] As a platform technology, the IC3D system can integrate other sensor methods (such as enzyme assays, PCR, and isothermal signal amplification) with droplet microfluidics and the 3D particle counter can be used for almost any type of low-abundance marker in biological samples. platform for the rapid detection and analysis of biological samples including cells (such as bacteria, circulating tumor cells, and stem cells), extracellular vesicles (such as exosomes), viruses (such as HIV), and molecular markers (such as nucleic acids and protein)( figure 1 ).

[0350] In alternative embodiments, the invention provides for the use of fluorescent substrates for beta-lactamases and carbapenemases for antimicrobial resistance IC3D tests, see e.g. Figure 26 . These tests enable the rapid detection of extended-spectrum beta-lactamase (ESBL) production in the most ubiquitous antimi...

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Abstract

In alternative embodiments, the invention provides high throughput, multiplexed systems or methods for detecting a biological, a physiological or a pathological maker, or a single molecule or a single cell using a droplet microfluidics system integrated with use of a sensor or a sensing system, an aptamer, or a DNAzyme. In alternative embodiments, the sensor or sensing system comprises a nucleic acid based, an antibody based, an enzyme based or a chemical based sensor or sensing system. In alternative embodiments, the invention provides methods for detecting a biological, a physiological or a pathological marker, or a single molecule or a single cell using a droplet system integrated with rapid and sensitive fluorescence detection systems including, for example, a 3D particle detector. In alternative embodiments, the invention provides systems comprising integrated comprehensive droplet digitial detection (IC 3D).

Description

technical field [0001] The present invention generally relates to biological assays, and methods of detection and screening. In particular, in alternative embodiments, the present invention utilizes integrated droplet microfluidic systems employing sensors or sensing systems, aptamers or DNAzymes, or emulsifiers to provide high-throughput, multiplexed systems or methods, For the detection of biological, physiological or pathological markers, or single molecules or single cells. In alternative embodiments, the sensor or sensing system comprises a nucleic acid based, antibody based, enzyme based or chemical based sensor or sensing system. In alternative embodiments, the present invention provides for the detection of biological, physiological or pathological markers using liquid droplet or emulsion systems integrated with rapid, sensitive fluorescent detection systems, including in particular 3D particle detectors, or single Molecular or single-cell approaches. In alternative...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K9/127C12M1/34C12M3/00
CPCC12Q1/6816G01N33/54366C12Q2525/205C12Q2537/143C12Q2563/159C12Q2565/607C12Q2565/629G01N33/56911G01N33/56916G01N33/57496
Inventor 赵伟安姜东求张开翔M·M·阿里M·A·埃克特李峰E·格拉顿M·A·迪格曼L·拉巴聂鲁梦柔
Owner RGT UNIV OF CALIFORNIA
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