الفهرس 
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Abstract
Vitrification is being introduced into clinical laboratories world-wide for human blastocyst preservation but it is still considered a new technology. However, ice crystal formation during the freezing process reduces the blastocyst survival rate. Artificial shrinkage or collapse of blastocysts prior to vitrification is a relatively new approach to improving blastocyst viability after vitrification. The theoretical basis for applying this technique prior to vitrifiation has been to help prevent ice crystal damage by reducing fluid within the blastocoel cavity.Laser technology is a simple, efficient, and precise cellular microsurgery tool that has been used in ART in recent years.
The aim of this study was to evaluate the potential effects of 1480 nm diode laser with different pulse duration, number of shots and hole sizeon the effectiveness of artificial shrinkage of blastocoeles before vitrification, on re-expansion and the survival rate after thawing.
In this study the following materials were used:
1- Laser pulse generated by a moveable Saturn 5 active laser (Bickland industrial park, Falmouth, cornwall TR114T.U.K.)
2- crytop (kitazato co, fujinomiya, Japan) was used to load blastocyst.
3- cryoprotectant solutions for vitrification and thawing procedures .
4- Human blastocysts:One hundred and thirty five blastocysts were divided into three main groups as follows:
- Control group: (45 Blastocyst) that is vitrified without reduction of blastocoelic fluid.
- Experimental group 1: (60 Blastocyst) Expanded blastocyst (4AA)
- Experimental group 2: (30 Blastocyst) early blastocyst (3AA)
Experimental group 1 and 2 were divided into 3 sub group according to number of shots, hole size, pulse width and time as follows:
• Sub group (a):
- No of shots fixed (one shot).
- Hole size/ pulse width was changed and time fixed at (2min) before vitrification.
• Sub group (b):
- No of shots fixed (one shot).
- Hole size/pulse width fixed at (0.697ms/9.8 µm) and time was changed.
• Sub group (c):
- No of shots changed.
- Hole size/pulse width fixed at (0.278ms/4.3 µm) and time fixed at (15 sec).
After application of artificial shrinkage of blastocyst with and without laser pulse, the following parameters were measured:
1- Length, width and zona thickness of blastocyst were measured before shrinkage and after warming.
2- Re-expansion was monitored 2 and 4 h after warming.
3- Survival rate was defined as the percentage of blastocysts with <50% degenerated on warming.
The experimental Results showed that:
• Hole size is directly proportional to pulse width, from linear relation increase in hole size could be calculated from equation Y= 0.0829X-0.1162.
• Expanded blastocyst artificially shrinkage seemed intact after thawing compared with blastocyst without artificial shrinkage.
• Immediately upon recovery, the laser collapsed blastocysts appeared to have the least fluid retention in their blastocoels cavity but also started to re-expand after two hours.
• The post-thawing re-expansion rate of zona treated with laser was significantly higher after four hours compared to re-expansion rate after two hours and the control group (p<0.05).
• High survival rates (55%) were obtained when blastocyst had been pretreated with different pulse width than untreated control group (40%).
• A significant increase in length of thawed blastocyst (101.82 vs 107.01) p <0.001) was observed in laser treated group when different times period were applied. Also, at just four hours, fully re-expanded blastocyst treated with laser was observed.
• High survival rate (70% vs 40%) was observed in laser treated group than control group respectively after applying different times for collapsing blastocele.
• Non-significant difference in survival rate between laser treated group and control group when multiple shots from laser pulse were directed to blastocyst.
• Survival rate of 3AA blastocysts with smaller blastocoelic volumes did not benefit from any further reduction of fluid volume.