Methods for Equine Preantral Follicles Isolation: Quantitative Aspects

Abstract

The aim of this study was to test the use of mechanical and mechanical-enzymatic methods, saline solution (SS), and PBS solution for the manipulation and isolation of mare ovarian preantral follicles (PAFs). The ovaries were subjected to mechanical isolation (mixer) alone or in association with enzymatic digestion (collagenase). Incubation times of 10 and 20 min were employed. In the first group, 4.1 ± 4.9 PAFs were harvested with the mechanical-enzymatic method vs 71.1 ± 19.2 with the mechanical procedure, showing a significant difference between methods; using SS and PBS, these numbers were 35.7 ± 34.3 and 39.6 ± 39.6, respectively, with no significant difference between solutions. In the second group, there was significant difference between methods, with 7.1 ± 10.6 follicles harvested with the mechanical-enzymatic method vs 63.2 ± 22.9 with the mechanical procedure; using SS and PBS, means were 35.5 ± 36.4 and 34.9 ± 31.1, respectively. The mechanical method proved more effective than the mechanical-enzymatic approach. Both SS and PBS can be used as a media for equine PAFs preparation.

Optimization of Porcine Ovarian Follicle Isolation Methods for Better Developmental Potential

“…Using hand blender. As the first attempt to isolate the porcine follicles with a mechanical method, we applied a hand blender (Bosch, Germany) as previously described by Nuttinck et al 27 and Leonel et al 28 In this method we put at least three ovaries into the hand blender container containing 4°C DPBS buffer. The ovaries were mixed until obtaining a homogenate of ovarian tissues.…”

Cryopreservation of Preantral Follicles

“…On the other hand, there are limitations, like follicle dissociation from the surrounding ovarian tissue. To isolate preantral follicles, we can use mechanical [69] or enzymatic [70] means, or an association of both, depending on the origin of the ovarian tissue [71]. The mechanical dissociation of the follicles is based on its fragmentation; cutting the cortex into small pieces with scissors or even with surgical blades are some options; in addition, the use of a tissue chopper has been widely described [71][72][73].…”