Bellio Pierangelo, Fagnani Lorenza, Nazzicone Lisaurora, Celenza Giuseppe
Department of Biotechnological and Applied Clinical Sciences, University of L'Aquila, L'Aquila, Italy.
MethodsX. 2021 Oct 11;8:101543. doi: 10.1016/j.mex.2021.101543. eCollection 2021.
To evaluate the effect of two combined antimicrobial drugs, one method currently in use is the checkerboard assay in a 96-well microplate, which gives a good estimation of the drug-drug combination effect. Appropriate and consolidated methods are described in numerous scientific publications which are, however, in turn, laborious and time-spending, specifically for the setting of the 96-well microplate preparation. Each drug of every combination must be prepared and dispensed individually in several steps, often limiting its use in terms of consumed materials and working time. In our method, the strengths of the previous consolidated techniques are kept, although the toughness and the execution time are drastically reduced. No special laboratory apparatuses are needed. All the procedures of our method can be referred to the CLSI or EUCAST guideline. The method provides few main steps, which can be summarised in:•Preparation of the microorganism inoculum and three concentrations of antimicrobial drugs.•Easy dispensing of all reagents into the microplates with a multichannel pipette.•Evaluation of the microorganism optical density (OD) by a microplate reader, and calculation of growth percentage for each of the 77 combinations.
为评估两种联合抗菌药物的效果,目前使用的一种方法是在96孔微孔板中进行棋盘法测定,该方法能很好地评估药物组合效果。众多科学出版物中描述了合适且完善的方法,然而,这些方法反过来又费力且耗时,特别是在96孔微孔板的制备方面。每种组合中的每种药物都必须经过几个步骤单独制备和分配,这往往在消耗材料和工作时间方面限制了其应用。在我们的方法中,保留了先前完善技术的优点,尽管难度和执行时间大幅减少。无需特殊的实验室设备。我们方法的所有程序都可参考CLSI或EUCAST指南。该方法提供了几个主要步骤,可总结如下:
• 制备微生物接种物和三种浓度的抗菌药物。
• 使用多通道移液器轻松将所有试剂分配到微孔板中。
• 通过微孔板读数器评估微生物光密度(OD),并计算77种组合中每种组合的生长百分比。