We developed a simple and fast microplate assay for evaluation of the antimicrobial activity of electrospun nanofiber filtration membranes or similar porous materials for water treatment technologies. Resazurin (alamarBlue^®) was used as an indicator of the amount of viable experimental microorganisms Gram-negative Escherichia coli, Gram-positive Enterococcus faecalis, and natural wastewater treatment plant effluent bacteria. A bacterial inoculum of concentration 1–3 × 10⁵ CFU mL⁻¹ was pipetted onto the surface of assessed both functionalized and respective control membranes and incubated in 12-well plates for 4 h at 37 °C. Kinetics of resazurin metabolization, i.e. its reduction to fluorescent resorufin, was evaluated fluorimetrically (λ_ex520/λ_em590 nm). A number of viable bacteria on the membranes expressed as CFU mL⁻¹ was calculated from the kinetic curves by using calibration curves that were constructed for both experimental bacterial species. Antimicrobial activities of the membranes were evaluated by either resazurin assay or modified ISO 20743 plate count assay. Results of both assays showed the significant antimicrobial activity of membranes functionalized with silver nanoparticles for both bacterial species and wastewater treatment plant effluent bacteria as well (log CFU reduction compared to control membrane > 4), while membranes containing specific quaternary ammonium salts were inefficient (log CFU reduction < 1). The suitability of resazurin microplate assay for testing nanofiber filtration membranes and analogous matrices has proven to be a faster and less demanding alternative to the traditionally used approach providing comparable results.

中文翻译：

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刃天青测定法评估电纺纳米纤维过滤膜的抗菌性能

我们开发了一种简单快速的微孔板测定法，用于评估用于水处理技术的电纺纳米纤维过滤膜或类似多孔材料的抗菌活性。刃天青 (alamarBlue ^{® ) 被用作活实验微生物（革兰氏阴性}大肠杆菌、革兰氏阳性粪肠球菌和天然废水处理厂废水细菌）数量的指示剂。将浓度为 1–3 × 10 ⁵ CFU mL ^-1的细菌接种物移至评估的功能化膜和各自的对照膜的表面上，并在 12 孔板中于 37 °C 下孵育 4 小时。通过荧光测定法评估刃天青代谢的动力学，即其还原为荧光试卤灵（λ _ex 520/λ _em 590 nm）。通过使用为两种实验细菌物种构建的校准曲线，根据动力学曲线计算膜上的活细菌数量，表示为CFU mL ^{-1 。}通过刃天青测定或改良的 ISO 20743 平板计数测定来评估膜的抗菌活性。两项测定的结果均表明，用银纳米颗粒功能化的膜对细菌种类和废水处理厂废水细菌均具有显着的抗菌活性（与对照膜相比，log CFU 减少> 4），而含有特定季铵盐的膜效率较低（log CFU 减少> 4） CFU 减少 < 1)。刃天青微孔板测定法适用于测试纳米纤维过滤膜和类似基质，已被证明是比传统使用的方法更快、要求更低的替代方法，可提供可比较的结果。