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Broth Microdilution Testing

Antimicrobial susceptibility testing (AST) is a critical part of antibacterial pharmacology and a basis for clinical medication. The importance of AST is to maintain the effectiveness of antimicrobial agents against epidemic pathogens, and to help select appropriate antimicrobial agents clinically. According to the guidelines of the Clinical and Laboratory Standards Institute (CLSI), AST is divided into dilution methods and disk diffusion methods. The broth microdilution testing belongs to dilution methods.

The broth microdilution testing is the most commonly used quantitative method to perform AST, which can economically, effectively, and accurately detect the minimal inhibitory concentration (MIC) of an antimicrobial agent with a pathogenic strain. Advantages of broth microdilution include: quantitative data is obtained to tell susceptibility or resistance of bacteria; multiple microtiter plates are commercially available, which supports standardized and harmonized testing; the testing system is easy to learn and to evaluate; and automation is possible.

Broth Microdilution Testing at Creative Animodel
Creative Animodel provides broth microdilution method to test the susceptibility of bacteria to antibiotics. In addition, the test can also be used to detect the efficacy of antimicrobial agents.

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Figure 1. Workflow of broth microdilution testing.

First of all, multiple microtiter plates are filled with Mueller-Hinton broth (MHB). Then, bacteria to be tested and varying concentrations of the antibiotics are added to the plate. The plate is then placed into a constant temperature incubator and heated at 35C for 16 to 24 hours. Following the incubation, the plate is removed and checked for bacterial growth. In general, observation with the naked eye can obtain the MIC directly (Figure 2). If the broth became cloudy or a layer of cells formed at the bottom, the bacterial growth has occurred. Besides, the MIC can also be determined by measuring the OD value of the broth using a microplate reader (Figure 3). According to the optical properties of bacteria, detection is usually performed at a wavelength of 600 nm. However, if the sample is not well-mixed for a long time, the bacteria will settle and the result will be affected. For colored drugs, the OD value will also be disturbed.

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Figure  2. Determine MIC by observing the turbidity directly. Figure  3. Determine MIC by measuring the OD value. (Prasad, T.; et al.  2006)

Creative Animodel has a team of experienced experts and advanced technology platforms in the field of AST and broth microdilution. We will deliver the detailed experimental report and accurate results in a time-saving manner. If you have any needs, please don’t hesitate to contact us.

Reference:
1. Prasad, T.; et al. Unexpected link between iron and drug resistance of Candida spp.: iron depletion enhances membrane fluidity and drug diffusion, leading to drug-susceptible cells. Antimicrobial Agents & Chemotherapy. 2006, 50(11):3597.

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