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Reagent, device and method for rapidly counting eukaryotic microorganisms

A microbial and eukaryotic technology, applied in the field of microbial detection, can solve the problems of reagents that cannot generate fluorescent signals in a short time, complex detection process, slow detection speed, etc., and achieve the effects of easy carrying and transportation, difficult diffusion, and easy identification

Pending Publication Date: 2022-03-04
ACAD OF NAT FOOD & STRATEGIC RESERVES ADMINISTRATION
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In order to solve the technical problems of the existing detection process, the dyeing reagent is absorbed and metabolized by destroying the microbial cell, the reagent that cannot generate a fluorescent signal in a short time, the detection speed is slow, the error is large, and the detection process is complicated, which is prone to cross-contamination

Method used

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  • Reagent, device and method for rapidly counting eukaryotic microorganisms
  • Reagent, device and method for rapidly counting eukaryotic microorganisms
  • Reagent, device and method for rapidly counting eukaryotic microorganisms

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0067] Embodiment 1, the reagent that is used for the fast counting of eukaryotic microorganism

[0068] This embodiment provides reagents for rapid counting of eukaryotic microorganisms, the reagents include osmosis buffer, fluorescent staining solution and reconstitution solution; wherein, the osmosis buffer includes the following components: ε-polylysine, dodecyl- β-D-maltoside, glycine.

[0069] Specifically, in the infiltration buffer, the contents of each component are: ε-polylysine 100 μmol / L, dodecyl-β-D-maltoside 0.2% by mass, and glycine 1 mmol / L;

[0070] Complex solution is dimethyl sulfoxide;

[0071] Fluorescent staining solution: 150 μg / mL 5-carboxyfluorescein diacetate, the preparation method of the fluorescent staining solution is to dissolve the fluorescent dyeing component 5-carboxyfluorescein diacetate in the complex solution dimethyl sulfoxide to form .

[0072] As an alternative embodiment, the components of the fluorescent staining solution can be sel...

Embodiment 2

[0073] Embodiment 2, the device that is used for the rapid counting of eukaryotic microorganism

[0074] The device for rapid counting of eukaryotic microorganisms in this embodiment includes a microorganism culture box and the reagent for rapid counting of eukaryotic microorganisms in Example 1.

[0075] like Figure 1A and Figure 1B As shown, the microbial cultivation box has a box body, the box body includes a holding tank 200 and a cover 100 matched with the holding tank, and the first absorbent paper 210, the second absorbent paper 210, and the second absorbent paper are sequentially laid in the holding tank 200 upwards from the bottom of the tank. The paper 220 and the filter membrane 230 ; the cover body 100 is provided with a pressing plate for pressing the first absorbent paper, the second absorbent paper and the filter membrane to the bottom of the holding tank.

[0076] Specifically, the microbial culture box can be a circular petri dish with a diameter of 50mm an...

Embodiment 3

[0077]Embodiment 3, the enumeration method to eukaryotic microorganism

[0078] The present embodiment provides the method for the enumeration of eukaryotic microorganism, it comprises the following steps:

[0079] Step 1, open the lid of the petri dish prepared in Example 2, add the sample to be tested containing yeast, the added volume of the sample is ≤5mL;

[0080] Add the dilution of the sample to be tested containing yeast to the center of the filter membrane, cover the petri dish, and remove the second absorbent paper after the dilution is completely expanded and absorbed;

[0081] Step 2, transfer the filter membrane with yeast absorbed to the first absorbent paper at the bottom of the culture dish, make the filter membrane adhere to the first absorbent paper, and ensure that there are no air bubbles between the two;

[0082] Step 3: Add 1.3mL liquid culture medium to the bottom of the petri dish, cover the petri dish, put it into an incubator at 28°C±1°C and incubate...

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Abstract

The invention discloses a reagent, a device and a method for rapidly counting eukaryotic microorganisms. The reagent comprises an osmotic buffer solution, a fluorescent staining solution and a reconstitution fluid, wherein the osmotic buffer solution is prepared from the following components: epsilon-polylysine, dodecyl-beta-D-maltoside and glycine; a staining component in the fluorescent staining solution is selected from fluorescein diacetate, 5 (6)-carboxyl-2 ', 5'-dihydroxyl-2 ', 5'-dihydroxyl-2 ', 5'- the fluorescein diacetate is one or more of 5, 7 '-dichlorofluorescein diacetate, 5-chloromethyl fluorescein diacetate, 5 (6)-carboxyl fluorescein diacetate, 6-carboxyl fluorescein diacetate, 5-carboxyl fluorescein diacetate, 5-nitryl-fluorescein diacetate and 6-nitryl-fluorescein diacetate. According to the reagent disclosed by the invention, a broad-spectrum bacteriostatic agent epsilon-polylysine is used as a carrier to transport a fluorescent staining solution to penetrate through cell walls and enter bacterial cell plasm, fluorescence is generated through excitation of exciting light, bacterial colonies are identified and counted, and a culture medium capable of accelerating the growth rate of thalli is used in a matched manner, so that the microbial detection efficiency is greatly improved.

Description

technical field [0001] The invention relates to the technical field of microorganism detection, in particular to a reagent, device and method for rapid counting of eukaryotic microorganisms. Background technique [0002] Microorganisms are small and diverse, including bacteria, fungi, viruses and other groups. Microbial colony is a group of cells that can be identified by us formed by the growth and reproduction of single or multiple microbial cells after cultured under suitable conditions. Microbial colony count monitoring analysis is widely used in many industries and is essential to ensure the quality and safety of products. [0003] In recent years, with the rapid development of the economy, people's living standards have been significantly improved, people's requirements for the quality of life are getting higher and higher, and the exposure rate of health issues such as food, medicine, cosmetics and the environment has increased significantly. Among them, mold and Ex...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/06C12M1/34C12M1/12
CPCC12Q1/06C12M41/36C12M25/02
Inventor 崔华王松雪王松山李森叶金郭宝元
Owner ACAD OF NAT FOOD & STRATEGIC RESERVES ADMINISTRATION
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