Figure 1

Schematic diagram for egg vitrification used in the study. A 20 μl of basic solution (BS) drop without CPAs and three 20 μl drops of equilibration solution (ES) are made in a cover of a 60 mm culture dish (1). Eggs are placed in the BS for 1 minute and then merged to 1 drop of ES using a transfer pipette for 2 minutes (2). The second ES drop is merged to the solution using the same transfer pipette for 2 minutes (3). Eggs are transferred from the merged solution to the third drop of ES for 5 minutes (4). When the eggs are in the third drop of ES, three 20 μl drops of vitrification solution (VS) are made in the same culture dish (4). Eggs are transferred to VS drop 1 (5), then to VS drop 2 (6), and finally to VS drop 3 (7). Eggs are remained in each drop for 10–20 seconds. The equilibrated eggs are finally loaded to the tip of the vitrification straw before the time reaches 90 seconds (8). Vitrification straw is inserted to cooled protective straw inside liquid nitrogen when the time reaches 90 seconds and the top end of the protective straw is sealed (9).