Enzyme linked immunosorbent assay instrument testing method for penicillium marneffei mycelia-phase in-vitro susceptibility testing

A Marneffei and Penicillium technology, applied in the field of Penicillium marneffei detection, can solve the problems of error in experimental results, difficulty in obtaining accurate results due to different personal judgments, and difficulty in judging breakpoints.

Inactive Publication Date: 2017-05-24
GUANGXI ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The in vitro susceptibility test of Penicillium marneffei is mainly based on CLSI M38-A 2 However, it is difficult to judge the breaking point because it uses the visual method to judge the results, and it is difficult to obtain accurate results due to different personal judgments, which brings certain errors to the experimental results.
Shi Luyi, Zhang Hanyuan and others from the Wildlife Research Institute of Heilongjiang Province used a microplate reader to interpret the results based on the microdilution method of the in vitro drug sensitivity test, which greatly avoided the lack of readings, but the method was only used in the large intestine. Bacillus, other strains have not been reported
Zhou Zixiong and others from the School of Traditional Chinese Medicine of Guangdong College of Pharmacy also conducted research on the determination of the antibacterial activity of antibacterial substances by microplate reader and established a method, but the method was only applied to Staphylococcus aureus

Method used

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  • Enzyme linked immunosorbent assay instrument testing method for penicillium marneffei mycelia-phase in-vitro susceptibility testing
  • Enzyme linked immunosorbent assay instrument testing method for penicillium marneffei mycelia-phase in-vitro susceptibility testing
  • Enzyme linked immunosorbent assay instrument testing method for penicillium marneffei mycelia-phase in-vitro susceptibility testing

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Prepare antibiotic liquid, accurately weigh 1.0mg of ketoconazole, dissolve it with 2mL dimethyl sulfoxide solvent to make antibiotic liquid;

[0021] Use RPMI 1640 culture medium to dilute the antibiotic solution to 4 μg / mL, then use a sterile microporous membrane (0.22 μm) to filter it for later use;

[0022]

[0023] Purification and cultivation of Penicillium marneffei: the lyophilized powder of Penicillium marneffei was cultivated and purified on PDA medium by dilution coating method;

[0024] Preparation of spore liquid: after the purified spores of Penicillium marneffei mature on the plate, draw 100-500 μL of RPMI 1640 culture solution with the tip of a sterile pipette and roll it on the plate to collect the spores. Collect into 10mL sterile plastic tube for use;

[0025] Cell counting of spore liquid: add the liquid in the plastic tube containing spores to 10mL, mix well, pipette 1-5μL liquid onto the hemocytometer plate, cover the glass slide and count und...

Embodiment 2

[0032] Prepare antibiotic liquid, accurately weigh 1.0mg of amphotericin B, and dissolve it with 2mL dimethyl sulfoxide solvent to make antibiotic liquid;

[0033] Use RPMI 1640 culture medium to dilute the antibiotic solution to 4 μg / mL, then use a sterile microporous membrane (0.22 μm) to filter it for later use;

[0034]

[0035] Purification and cultivation of Penicillium marneffei: the lyophilized powder of Penicillium marneffei was cultivated and purified on PDA medium by dilution coating method;

[0036] Preparation of spore liquid: after the purified spores of Penicillium marneffei mature on the plate, draw 100-500 μL of RPMI 1640 culture solution with the tip of a sterile pipette and roll it on the plate to collect the spores. Collect into 10mL sterile plastic tube for use;

[0037] Cell counting of spore liquid: add the liquid in the plastic tube containing spores to 10mL, mix well, pipette 1-5μL liquid onto the hemocytometer plate, cover the glass slide and cou...

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Abstract

The invention discloses an enzyme linked immunosorbent assay instrument testing method for penicillium marneffei mycelia-phase in-vitro susceptibility testing. The light transmittance of a bacterium-containing diluent with an antibiotic medicinal liquid is tested by using an enzyme linked immunosorbent assay instrument in the penicillium marneffei mycelia-phase in-vitro susceptibility testing process. The densities are different in different bacterium growth situations, the light transmittances are also different, results are judged by using the enzyme linked immunosorbent assay instrument, apparent phenomena can be converted into values, and the operation can be relatively convenient and accurate; secondly, as an enzyme linked immunosorbent assay instrument pore plate is used, material consumption can be reduced; only a solution less than 1ml is needed in each pore, so that the amount of medicines is small, and waste can be greatly reduced; in addition, penicillium marneffei has infectivity, so that the pollution property can be further reduced because of the small use amount. Finally, normal growth of the penicillium marneffei can be not affected by the enzyme linked immunosorbent assay instrument, the operation is simple, convenient and rapid, and precise in-vitro susceptibility data results can be obtained.

Description

technical field [0001] The invention belongs to the technical field of Penicillium marneffei detection, and in particular relates to an enzyme-labeled detection method for an in vitro drug susceptibility test of Penicillium marneffei hyphae. Background technique [0002] Penicillium marneffei infection is the main infection factor leading to the death of AIDS patients in southern China, especially in Guangxi and Guangdong. Penicillium marneffei can cause deep fungal infection when human immunity is low, and invade multiple organs throughout the body. If it is not treated in time and correctly, it can lead to death. At present, the main therapeutic drugs for Penicillium marneffei are antibiotics, and amphotericin B (AmB), fluconazole, flucytosine, etc. are commonly used drugs for the treatment of Penicillium marneffei. [0003] Penicillium marneffei is the only pathogenic bacterium in Penicillium that exhibits a temperature biphasic type. It is in a mycelial state at a tempe...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/04C12R1/80
CPCC12Q1/04
Inventor 高程海李家怡李菲何碧娟周文红余炼劳超
Owner GUANGXI ACAD OF SCI
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