THE EFFECTIVENESS OF CRYOPROTECTANT AND Ca2+ DISTRIBUTION ON SPERM VITRIFICATION

• Main Author: Widyastuti, Rini
• Format: Theses
• Language: Indonesia
• Subjects: Ilmu hayati ; Biologi
• Online Access: https://digilib.itb.ac.id/gdl/view/36109
• Institution: Institut Teknologi Bandung

Vitrification is one of methods that could be used for sperm cryopreservation. The aim of this study was to observe the effect of seminal plasma to sperm recovery rate, the effectiveness of cryoprotectants and Ca2+ distribution after vitrification process. Twenty-four ejaculated with concentration 20 million/ml, progressive motility and viability above 50% were used as samples. In Experiment 1, samples were divided into 2 groups, i.e with and without seminal plasma. We were using modified EBSS medium contain 0.25 M sucrose, 1% HAS and 36.25% EG as vitrification medium. Sperm packaged in 0.25 ml straws were equilibrated for 10 minutes before plunged into liquid nitrogen. Samples were warmed after 24 hours for sperm analysis. The result showed that the recovery rates were lower in the group without plasma than the other groups. In Experiment II, we were using different concentration (36,25%; 18,25%; 9,12%; 4,56%; 1,14% and 0,57%) of EG in the EBSS + 1% HAS medium. The results showed that the lowest concentration of EG produced the highest sperm recovery rate after warming. In Experiment III, we were using 3 groups of medium; a) EBSS medium with 0,25 M sucrose, 1% HAS and 0.57% EG, b) EBSS medium with 0.25 M sucrose and 1% HAS and c) EBSS medium without cryoprotectant. The results showed that the highest although no statistic significant different (P<0.05) recovery rate was in EBSS medium with 0.25 M sucrose and 1% HAS (b). In Experiment IV, we observed the effect of vitrification to the sperm using Ca Fluro-3 as an indicator. The distribution of Ca2+ were significantly different between motile and immotile sperm after vitrification process. In motile sperms, the Ca2+ was spread all part of the sperm head, but in immotile sperm the Ca2+ was accumulated in the posterior part of the head and anterior part of the midpiece.