1.1 本实践提供了在平面上创建细菌细胞单层的协议。 1.2 本规程中使用的培养物和培养物制备步骤类似于AOAC方法961.02和美国EPA MB-06。然而，使用自动沉积设备将测试细菌应用于载体( 6.2 )而不是悬浮液滴。 1.3 载体检查协议类似于美国EPA MB-03，不同之处在于载体表面是通过显微镜检查而不是通过肉眼进行检查。 1.4 单层细胞消除了由细菌外层叠加在试样上其他细菌上的阴影效应引起的混淆效应，从而衰减了定向能束（即紫外线、高强度辐射）- 在电子束到达底层细胞之前。 1.5 无凹凸表面消除了样本表面拓扑结构的阴影效应，该拓扑结构可以阻止目标细菌直接暴露于非化学抗菌处理。 1.6 本规程提供了可复制的目标微生物和表面样本，以最小化测试设施内和测试设施之间的样本可变性。这有助于在各种非化学抗菌技术之间进行直接数据比较。 1.6.1 临床和工业应用中使用的抗菌杀虫剂有望克服阴影效应。然而，这种做法满足了对协议的需要，该协议有助于在非- 化学抗菌处理。 1.6.2 本规程不旨在满足或取代液体化学抗菌处理的现有试验要求（例如试验方法 E1153 和 E2197年 )或制定监管机构绩效标准，如美国环保局MB-06。 1.7 该实践通过使用 金黄色葡萄球菌 （ATCC 6538）和 铜绿假单胞菌 （ATCC 15442）使用基于AOAC方法961.02的协议。如果使用其他培养物，则必须通过使用扫描电子显微镜（SEM）或类似的高分辨率显微镜检查制备的表面来确认此做法的适用性。 1.8 根据本规程制备的试样不用于模拟端部- 使用条件。 1.8.1 非化学技术只能用于明显清洁、无孔的表面。因此，不使用土壤荷载。 1.9 单位- 以国际单位制表示的数值应视为标准值。本标准不包括其他计量单位。 1.10 本标准并非旨在解决与其使用相关的所有安全问题（如有）。本标准的用户有责任在使用前制定适当的安全、健康和环境实践，并确定监管限制的适用性。 1.11 本国际标准是根据世界贸易组织技术性贸易壁垒（TBT）委员会发布的《关于制定国际标准、指南和建议的原则的决定》中确立的国际公认标准化原则制定的。 ====意义和用途====== 5.1 没有可复制的标准化方案用于制备用于评估非化学处理（如紫外线、高能电子束或其他形式的非化学抗菌技术）的杀菌效果的样本。 5.2 将生物负载应用于载体的常规协议（见测试方法 E2197年 )使细胞相互堆叠，从而形成多个细胞层，其中靠近载体的层中的细胞被覆盖层中的细胞遮盖，这使得不同非化学抗菌处理的相对比较更加困难。 5.3 钢和其他金属载体具有微凸体，可以屏蔽一定比例的应用电池，使其不直接暴露于电磁辐射。 5.4 的综合影响 5.2 和 5.3 电磁辐射对试样杀菌效果的混杂测定。 5.5 该实践通过以下方式解决了这两个混杂因素: 5.5.1 使用玻璃显微镜载玻片（其表面无粗糙度）作为载体。 5.5.2 将细菌细胞作为单层可靠地沉积在载体上。 5.6 由此产生的样本确保沉积在载体上的所有微生物平等地暴露在辐射源中，从而确保唯一的变量是受控变量-起始接种物浓度、波长（λ–nm）、暴露时间（s）和产生的能量剂量（J）。

1.1 This practice provides a protocol for creating bacterial cell monolayers on a flat surface. 1.2 The cultures used and culture preparation steps in this Practice are similar to AOAC Method 961.02 and US EPA MB-06. However, test bacteria are applied to the carrier using an automated deposition device ( 6.2 ) rather than as a suspension droplet. 1.3 The carrier inspection protocol is similar to US EPA MB-03 except that carrier surfaces are inspected microscopically rather than visually, unaided. 1.4 A monolayer of cells eliminates the confounding effect caused by the shadowing effect of outer layers of bacteria stacked upon other bacteria on test specimens – thereby attenuating directed energy beams (that is, ultraviolet light, high-energy electron beams) before they can reach underlying cells. 1.5 An asperity-free surface eliminates the shadowing effect of specimen surface topology that can block direct exposure of target bacteria to non-chemical antimicrobial treatments. 1.6 This practice provides a reproducible target microbe and surface specimen to minimize specimen variability within and between testing facilities. This facilitates direct data comparisons among various non-chemical antimicrobial technologies. 1.6.1 Antimicrobial pesticides used in clinical and industrial applications are expected to overcome shadowing effects. However, this practice meets a need for a protocol that facilitates relative comparisons among non-chemical antimicrobial treatments. 1.6.2 This practice is not intended to satisfy or replace existing test requirements for liquid chemical antimicrobial treatments (for example Test Methods E1153 and E2197 ) or established regulatory agency performance standards such as US EPA MB-06. 1.7 This practice was validated using Staphylococcus aureus (ATCC 6538) and Pseudomonas aeruginosa (ATCC 15442) using a protocol based on AOAC Method 961.02. If other cultures are used, the suitability of this practice must be confirmed by inspecting prepared surfaces, by using scanning electron microscopy (SEM) or comparable high-resolution microscopy. 1.8 The specimens prepared in accordance with this practice are not meant to simulate end-use conditions. 1.8.1 Non-chemical technologies are only to be used on visibly clean, non-porous surfaces. Consequently, a soil load is not used. 1.9 Units— The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard. 1.10 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use. 1.11 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee. ====== Significance And Use ====== 5.1 There are no reproducible standardized protocols for preparing specimens used to evaluate the microbicidal efficacy of non-chemical treatments such as ultraviolet (UV), highenergy electron beam, or other forms of non-chemical antimicrobial technologies. 5.2 Conventional protocols for applying bioburdens to carriers (see Test Method E2197 ) cause cells to stack upon one another, thereby creating multiple cell layers in which cells in layers closer to the carrier are masked by cells in overlying layers, which makes relative comparison of different non-chemical antimicrobial treatments more difficult. 5.3 Steel and other metal carriers have asperities that can shield a percentage of the applied cells from direct exposure to electromagnetic irradiation. 5.4 The combined effects of 5.2 and 5.3 confound determination of the microbicidal effect of electromagnetic irradiation on test specimens. 5.5 The practice addresses these two confounding factors by: 5.5.1 Using glass microscope slides – the surfaces of which are asperity-free – as carriers. 5.5.2 Reliably depositing bacterial cells onto the carrier as a monolayer. 5.6 The resulting specimen ensures that all microbes deposited onto the carrier are exposed equally to the irradiation source thereby ensuring that the only variables are the controlled ones – starting inoculum concentration, wavelength (λ – in nm), exposure time(s), and resulting energy dose (J).