HOW TO OPTIMIZE CULTURE MEDIA OSMOLALITY IN IVF TREATMENTS

Abstract

Study aim was compare osmolality in sequential and single step culture media, covered with two types of oil mineral and paraffin, in two types of incubators, dry and humid. A prospective observational study was performed from March to November 2019. For each incubator, humid water jacket (Forma 4130, Thermo Scientific) and dry bench top (G185, K-Systems), both trigas, a total of 30 dishes were prepared with sequential (Cleavage Medium, Cook) and single step media (CSCM, Irvine Scientific), covered with paraffin oil (Ovoil, Vitrolife). Another 30 dishes were prepared as mentioned above, covered with mineral oil (Light Mineral Oil, Irvine Scientific). Statistical analysis was performed using ANOVA. Twenty 35x10 millimeters dishes (Falcon), at a time, were prepared in a laminar flow hood at room temperature, using 25 microliter droplets of CSCM and Cleavage Medium. Ten were covered with 3,0 mL mineral oil and ten with paraffin oil. Half of the dishes were incubated in a dry incubator and another half in a humid. Osmolality (mOsm/kg) was measured using osmometer (Advanced Instruments) on day 1, 3, 5 and 7, one drop per day. Humid incubator is better for maintaining the osmolality of culture media, regardless of the type of oil used. CSCM has lowest levels of osmolality on day 1 when compared to Cleavage (263 X 286, p<0.0001) and maintained statistically lowest levels of osmolality on day 3, 5 and 7. In humid incubator, both culture media using paraffin or mineral oil, had similar osmolality on day 3 (269 X 270, P=0.391 for CSCM; 293 X 292, P=0.759 for Cleavage) , day 5 (270 X 271 P=0.978 for CSCM; 292 X 293 P=0.914 for Cleavage) and day 7 (271 X 271 P=0.999 for CSCM; 294 X 292 P=0.624 for Cleavage). Paraffin oil provided greater protection against evaporation, maintaining lower levels of osmolality, in a dry incubator, for CSCM media in D3 (281 x 284, P<0.001), D5 (287 x 293, P<0.001) e D7 (298 x 304, P<0.001). No significant difference was observed for Cleavage osmolality on day 3 and 5 (304 X 306, P=0.213) (314 X 314, P=1.0) comparing two types of oil, in a dry incubator. On day 7 paraffin oil is better to maintain osmolality (322 X 325, P=0.003). Our study demonstrated that combination of humid incubator and paraffin oil are better for maintaining osmolality level of single step media, which can improve continuous and undisturbed embryonic culture. Comparing humid versus dry benchtop incubators, using different types of oils and culture media, is necessary for better IVF results.