Microdetermination method for measuring minimum inhibitory concentration and minimal bactericidal concentration of cationic antimicrobial peptide

A technology of antibacterial concentration and bactericidal concentration, which is applied in the direction of biochemical equipment and methods, microbial measurement/inspection, etc., can solve the problems of unsatisfactory demand, detection impact, and few antibacterial peptides, and achieve economical reagents and easy operation , the effect of simple equipment

Active Publication Date: 2013-06-26
ZHEJIANG UNIV
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Problems solved by technology

[0003] Since cationic antimicrobial peptides carry positive charges, the concentration of cations in the antibiotic-sensitive assay medium used in general detection methods and the adsorption of the utensils used will have a greater impact on the detection
In addition, the amount of samples to be tested for general antimicrobial peptides is very small, which cannot meet the requirements of general detection methods

Method used

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  • Microdetermination method for measuring minimum inhibitory concentration and minimal bactericidal concentration of cationic antimicrobial peptide
  • Microdetermination method for measuring minimum inhibitory concentration and minimal bactericidal concentration of cationic antimicrobial peptide

Examples

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Embodiment 1

[0041] On September 18, 2012, the minimum inhibitory concentration and minimum bactericidal concentration of antibacterial peptide PR-39 and antibiotics aureomycin, neomycin and colistin sulfate on Escherichia coli CMCC 44103 and ATCC 25922 were determined.

[0042] ) Reagent preparation

[0043] ①MHB medium: 21 g of MHB medium powder (BD Difco, product number 275730) was dissolved in 1 L of distilled water, sterilized at 121 °C for 20 min, cooled at room temperature, and stored at 4 °C for later use.

[0044] ②MHB agar medium: Dissolve 21 g of MHB medium powder and 15 g of agar powder in 1 L of distilled water, sterilize at 121 °C for 20 min, pour into a sterile petri dish, preferably just covering the bottom of the petri dish, cool and solidify at room temperature , stored at 4°C for later use.

[0045] ③Medicine storage solution: Accurately weigh 2 mg of the drug to be tested, dissolve it in sterile water to 4 mg / ml, and store it at 4 °C for later use.

[0046] ) Bac...

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Abstract

The invention discloses a microdetermination method for measuring minimum inhibitory concentration and minimal bactericidal concentration of cationic antimicrobial peptide. The method comprises the following steps: 1) preparing a reagent, namely preparing an MHB culture medium; 2) preparing bacterial suspension, namely reviving the bacteria to logarithmic phase, and preparing bacterial suspension with a certain concentration; 3) counting the bacterial suspension, namely performing coubling dilution on the bacterial suspension in the step 2), dripping and counting; 4) diluting the agent, namely performing coubling dilution on the agent to different testing concentration gradient; 4) measuring the minimum inhibitory concentration, namely mixing and culturing the diluted agent in the step 4) with the bacterial suspension prepared in the step 2), and observing whether macroscopic bacterial precipitation exists; and 6) measuring the minimal bactericidal concentration, namely culturing the testing dripping plate with the antibacterial effect in the step 5), and observing whether colony growth exists. The method for measuring the minimum inhibitory concentration and minimal bactericidal concentration of cationic antimicrobial peptide has the characteristics of low interference, trace, convenience and rapidness.

Description

technical field [0001] The invention belongs to a method for measuring the minimum inhibitory concentration and the minimum bactericidal concentration of trace antibacterial substances, in particular suitable for the determination of cationic antibacterial peptides. Background technique [0002] Antimicrobial peptides are a large class of biologically active substances with antibacterial activity and immune function in animals. Antimicrobial peptides have broad-spectrum antibacterial activity and rich mechanisms of action. Not only the same as traditional antibacterial active substances, but also has intracellular mechanism of action, including: binding to nucleic acid substances, blocking DNA replication and RNA synthesis; affecting protein synthesis; inhibiting membrane and cell wall synthesis, hindering cell division; inhibiting the activity of intracellular enzymes. Antimicrobial peptides, especially cationic antimicrobial peptides, have a membrane action mechanism. Cur...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/18
Inventor 汪以真李治学韩菲菲黄霞刘丹
Owner ZHEJIANG UNIV
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