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System for rapidly, directly, absolutely and quantitatively detecting microbes based on liquid drop, and method thereof

An absolute quantitative and detection system technology, applied in biochemical equipment and methods, microbial measurement/inspection, enzymology/microbiology devices, etc., can solve the problem of low efficiency, cumbersome monitoring process, unsuitable for real-time on-site or process detection, and cultivation cycle Long-term problems, to achieve the effect of low energy consumption

Inactive Publication Date: 2015-09-02
QINGDAO INST OF BIOENERGY & BIOPROCESS TECH CHINESE ACADEMY OF SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the traditional plate-based culture detection method has a long culture cycle, low efficiency, and cumbersome monitoring process, which is not suitable for instant on-site or process detection.
Other microbial detection methods including immunity, nucleic acid amplification, ATP luminescence and other microbial detection methods have a short analysis cycle and are suitable for rapid detection. However, these methods are relatively quantitative methods with high false positive or false negative detection rates and limited accuracy. Use culture methods to confirm assay results

Method used

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  • System for rapidly, directly, absolutely and quantitatively detecting microbes based on liquid drop, and method thereof
  • System for rapidly, directly, absolutely and quantitatively detecting microbes based on liquid drop, and method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] Example 1: Portable microbial detection based on droplet technology

[0040] like figure 1 As shown, in the portable microbial detection system and method based on droplet technology constructed in this example, the microfluidic chip 9 includes a sample suspension inlet 1 containing a culture medium, an oil phase injection port 2 containing a surfactant, Droplet generation channel 3 , droplet waste port 5 . The chip consists of two layers of PDMS, the upper layer includes a microchannel network with a width of 25-100um and a depth of 30-70um, and the lower layer is a PDMS substrate. The upper and lower substrates are cleaned and then bonded using plasma. Vacuum the chip with negative pressure, or use micro-syringe negative pressure suction to generate driving force. After the sample is subjected to microbial enrichment pretreatment, etc., 1 ml of suspension (containing microbial culture medium and color indicator of cell metabolites) is formed, and an appropriate con...

Embodiment 2

[0041] Embodiment 2: laboratory microorganism detection based on droplet technology

[0042] like figure 1 , 2 As shown, the microfluidic chip 9 constructed in this example is used for mutant screening based on droplet single-cell encapsulation under flow conditions, and the chip includes a sample suspension inlet 1 containing a culture medium, an oil containing a surfactant. Phase injection port 2 , droplet generating channel 3 , droplet waste liquid port 5 . The microfluidic chip 9 consists of two layers of PDMS, the upper layer includes a microchannel network with a width of 25-100um and a depth of 30-70um, and the lower layer is a PDMS substrate. The upper and lower substrates are cleaned and then bonded using plasma. The inlet and outlet of the chip are connected with the outside world through fixed interfaces and microtubes to realize the introduction and separation of liquid flow. After the sample is subjected to microbial enrichment pretreatment, etc., a 1 ml suspe...

Embodiment 3

[0043] Example 3: Online microbial index detection of cleaning products based on droplet technology

[0044]In this example, the constructed droplet microbial detection system and method are applied to the online microbial quality control detection of cleaning products, and the system and method described in Example 2 are adopted. Cleaning products are enriched by microbial membrane filtration, suspended in culture medium to form a microbial sample suspension, and an appropriate amount of fluorescent dye alarmblue is added, and an appropriate concentration of surfactant is added to the oil phase to push them into the microchannel network of the chip with a driving force. The fluid flows in an orderly manner in the closed microchannel network to form monodisperse microbial cell-wrapped droplets, which are collected into the culture detection pool and cultured under temperature control. Fluorescence microscopes are used to magnify the droplets in the detection area to collect bri...

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Abstract

The invention relates to a system for rapidly, directly, absolutely and quantitatively detecting microbes based on a liquid drop by using a liquid drop micro-fluidic technology, and a method thereof. The system comprises a micro-fluidic chip, a plurality of cell collecting, culturing and observing chambers, an optical detection unit and an image processing unit. The method comprises the following steps: preprocessing a sample, injecting the preprocessed sample into the micro-fluidic chip to form monodispersed unicellular liquid drops, and carrying out controlled temperature cell culture on the monodispersed unicellular liquid drops formed in the cell collecting, culturing and observing chambers for 1-2h; and determining the existence of living cells and the growth speed of the cells according to the turbidity of a single liquid drop, collecting the amplified pictures of the liquid drops in the cell collecting, culturing and observing chambers, and counting the turbid liquid drops to obtain the quantity of the microbes in the sample. The system for rapidly, directly, absolutely and quantitatively detecting microbes based on a liquid drop and the method have the advantages of simplicity and low cost, are suitable for portable onsite detection and laboratory detection, and can be used to detect microbe indexes in the fields of foods, environment, drugs sanitation products and cosmetics.

Description

technical field [0001] The present invention constructs a rapid microbial direct absolute quantitative detection system and method based on droplet single-cell encapsulation technology, and provides a rapid detection system and method for rapid detection of microorganisms in food, medicine, cosmetics, cleaning products, etc., the present invention It belongs to the field of biotechnology and rapid detection. Background technique [0002] Microbial pollution exists in every link of the whole process of production, processing, storage, transportation, sales, and use of food, cosmetics, and Procter & Gamble products. few key links. The culture-based microbial detection method has the characteristics of high accuracy and is often used as the gold standard and confirmation method for microbial detection of food, medicine and cosmetics. However, the traditional plate-based culture detection method has a long culture cycle, low efficiency, and cumbersome monitoring process, which...

Claims

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Application Information

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IPC IPC(8): C12M1/34C12Q1/06
Inventor 马波张旭徐健
Owner QINGDAO INST OF BIOENERGY & BIOPROCESS TECH CHINESE ACADEMY OF SCI
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