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Separation method of rickettsia intracellular bacteria strain

A technology for rickettsia and strain separation, applied in the field of medical microbiology, can solve the problems of difficulty in strain separation, difficulty in culturing, and low purity of strains, and achieve the effect of simple, easy-to-operate and far-reaching separation methods.

Pending Publication Date: 2021-08-10
李家斌
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] The purpose of the present invention is to solve the problems in the prior art that the isolation of Rickettsia intracellular bacteria strains is relatively difficult, the isolated bacterial strains have low purity, and it is difficult to cultivate them in cells, and a kind of Rickettsia intracellular bacteria proposed Strain Isolation Method

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  • Separation method of rickettsia intracellular bacteria strain

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Embodiment 1

[0028] A kind of Rickettsia intracellular bacterial strain isolation method that the present invention proposes comprises the following steps:

[0029] (1) Through preliminary experiments, it was found that Rickettsia grew well in mouse fibroblasts (L929 cells), so the bacterial fluid or serum containing Rickettsia was infected into L929 cells, and after 3 days of infection, Smear the cells every day and perform Giemsa staining, observe the number of bacterial cells in the cytoplasm under a microscope, until the cytoplasm is full of bacterial cells, all the adherent cells infected with Rickettsia can be scraped off for later use; Kleutzia is an intracellular parasite, which grows in the cytoplasm of the L929 cells it infects, and Giemsa staining shows purple short rods;

[0030] (2) Add the adherent cells with Rickettsia infection scraped off in step (1) and the cell culture solution obtained by compounding DMEM medium, HEPEs buffer and glutamine into 50ml centrifuge tube A ,...

Embodiment 2

[0043] A kind of Rickettsia intracellular bacterial strain isolation method that the present invention proposes comprises the following steps:

[0044] (1) Through preliminary experiments, it was found that Rickettsia grew well in mouse fibroblasts (L929 cells), so the bacterial fluid or serum containing Rickettsia was infected into L929 cells, and after 3 days of infection, Smear the cells every day and perform Giemsa staining, observe the number of bacterial cells in the cytoplasm under a microscope, until the cytoplasm is full of bacterial cells, all the adherent cells infected with Rickettsia can be scraped off for later use; Kleutzia is an intracellular parasite, which grows in the cytoplasm of the L929 cells it infects, and Giemsa staining shows purple short rods;

[0045] (2) Add the adherent cells with Rickettsia infection scraped off in step (1) and the cell culture solution obtained by compounding DMEM medium, HEPEs buffer and glutamine into 50ml centrifuge tube A ,...

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Abstract

The invention belongs to the technical field of medical microbiology, particularly relates to a separation method of a rickettsia intracellular bacterium strain, and solves the problems that in the prior art, the rickettsia intracellular bacterium strain is difficult to separate, and the separated strain is low in purity and is difficult to culture in cells. The preparation method of the separation method of the rickettsia intracellular bacterium strain comprises the following steps: centrifuging, crushing and centrifuging adherent cells infected by rickettsia to obtain a primary pure strain, adding meglumine diatrizoate for ultracentrifugation to obtain an intra-layer bacterium suspension, and finally centrifuging and fixing the volume to obtain the rickettsia intracellular bacterium strain. The separation method of the rickettsia intracellular bacteria strain is simple and easy to operate, the rickettsia intracellular bacteria are separated from cells, the rickettsia intracellular bacteria strain with high purity is obtained, the method can be used for subsequent research on the pathogenic mechanism and specific clinical manifestation of rickettsia, and the significance is profound.

Description

technical field [0001] The invention relates to the technical field of medical microbiology, in particular to a method for isolating Rickettsia intracellular bacterial strains. Background technique [0002] Rickettsia generally refers to Rickettsia, which is a Gram-negative bacterium and a class of G-prokaryotes obligately parasitic in eukaryotic cells. It is a kind of prokaryote close to bacteria, between bacteria and viruses, without nucleolus and nuclear membrane. Generally spherical or rod-shaped, it mainly parasitizes arthropods, and some of them can be transmitted to the human body through fleas, lice, ticks, and mites, causing typhus, trench fever and other diseases. Rickettsia was first discovered and reported by the young doctor Howard Taylor ricketts in 1906, but he was unfortunately infected and died in the process of researching Rickettsia. To commemorate his dedication in research and the discovery of Rickettsia He has made outstanding contributions, so this t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/02C12R1/01
CPCC12N1/02
Inventor 温萌李家斌殷俊程君叶英陈振胡立芬刘艳艳柳燕马雪娇常啸
Owner 李家斌
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