Parameters of the Mouse Embryo Assay that affect detection of peroxides in mineral oil

Abstract

Research question: Can culture conditions influence the sensitivity of a Mouse Embryo Assay and its potential to detect peroxide-related toxicity in mineral oil samples?

Design: Protein type and concentration, embryo density and culture dish design were selected as the variables in the culture system with the potential to influence the assay's sensitivity. Fresh 1-cell mouse embryos were cultured under mineral oil samples with known peroxide concentrations. Protein type (human serum albumin [HSA] + α/β-Globulins versus HSA versus bovine serum albumin [BSA]), concentration (5 mg/ml versus 0.5 mg/ml), embryo density (25 versus 3 µl/embryo) and culture dish (Petri versus micro-well dish) were adjusted to define the culture conditions with the highest sensitivity.

Results: High concentrations of peroxides can be easily detected by current quality control standards. However, for oil samples with a lower concentration of peroxides, supplementing the culture medium with 5 mg/ml of HSA + alpha/beta-globulins or with HSA resulted in an increased detection of embryo toxicity compared with when BSA was used as the protein supplement. The sensitivity of the assay was greatly reduced when embryos were cultured in groups and when certain micro-well dishes were used.

Conclusions: Current quality control protocols may not be sensitive enough to identify low concentrations of peroxides, which, if undetected, can increase over time and become potentially harmful during gamete and embryo culture. The different parameters established in this study allow the sensitivity of the Mouse Embryo Assays to be optimized to specifically detect peroxides in mineral oil samples prior to their release into the market and their broad use in human IVF.

Keywords: Mineral oil; Mouse Embryo Assay; Peroxides; Quality Control.