The micro‐Ames test: A direct comparison of the performance and sensitivities of the standard and 24‐well plate versions of the bacterial mutation test

Abstract

"Ames" bacterial mutation tests are widely performed for evaluation and registration of new materials including industrial chemicals, agrochemicals, medical devices, pharmaceuticals, pharmaceutical impurities and other materials. Tests are used to predict their potential long-term adverse health effects (including carcinogenicity). Given their importance, pre-screening 'miniaturized' versions have been developed which allow higher throughput and use less test material, including the widely-employed 24-well micro-Ames (µAmes) test which uses 20 times less material. However, little quantitative information has been published on the methodology or sensitivity of this system. We describe methods and results used in direct comparisons of the sensitivity of micro and standard systems using the same cultures, formulations, etc. Initial testing utilized the plate incorporation method and, later, the pre-incubation method. In a subsequent phase of testing, a four-way direct comparison was made between the pre-incubation and plate incorporation methods in both systems using some direct-acting mutagens. Tests used only those strain/S9/chemical combinations where a response was expected. Historical control results accumulated during testing are also presented. Spontaneous and induced revertant colony counts for the µAmes system were consistently proportionate and approximately 1/20th those for the standard Ames test. Sensitivities of the two systems were found to be nearly identical in almost all cases for a wide variety of weak and strong inorganic and organic mutagens. Standardized procedures and increased reliability of the estimate of the background revertant frequency in the µAmes system means that the two systems give equivalent results and are expected to be highly predictive of one another. Environ. Mol. Mutagen. 57:687-705, 2016. © 2016 Wiley Periodicals, Inc.

A novel small molecule chaperone of rod opsin and its potential therapy for retinal degeneration

“…To assess the risk of tumorgenesis by YC-001 treatment, Ames bacterial mutation tests were performed in a total of five bacteria strains 45 . The negative results of YC-001 in the Ames test (Supplementary Table 3 ) suggests that YC-001 has a low risk of mutagenicity if tested in vivo.…”

Comparação do perfil de mutagenicidade e da composição quí­mica do material particulado atmosférico de Limeira, Estocolmo e Quioto

“…Another advantage of the MPA is anticipated in EDA studies where several fractions need to be tested with the objective to find the most potent ones to perform chemical characterizations (Brack et al 2016). The MPA protocol could also be applied in specific situations such as drug impurity testing when only few mg of sample are available, with the limitations already pointed out by Proudlock and Evans (2016). The MPA protocol allows for the application of more strains with the same amount of sample used in the microsuspension protocol, enhancing the ability of detection of a mutagenic response for chemicals like C.I.…”

“…We decided to use only one tube to reduce as much as possible the quantity of sample used in the assay. This test design was based on the miniaturized preincubation protocol described by Proudlock and Evans (2016) except that they used 3 wells derived from a single treatment and we used 4 wells.…”

“…The Salmonella/microsome assay is the most widely used mutagenicity test both for evaluation of pure chemicals and environmental samples and is considered a reliable tool in the toxicology of the 21st century . Some researchers have modified the test conditions and developed reduced volume protocols that are available in the literature Brooks 1995;Flamand et al 2001;Pant et al 2016;Proudlock and Evans 2016). These modifications provide clear advantages, such as cost-effectiveness, reduced hands-on time and especially the reduction in S9 mix and sample volume needed.…”

“…Several other reduced protocols using agar media have been developed (Brooks 1995;Flamand et al 2001) which used 2.5 times less sample and 5 times less S9 mix per plate in comparison with the standard protocol . Pant et al (2016) used 5 times less sample and Proudlock and Evans (2016) used 20 times less sample and S9 per plate. All agar-based reduced protocols provided good agreement with the standard protocol.…”