Freezing goat embryos at different developmental stages and quality using ethylene glycol and a slow cooling rate

Abstract

The efficiency of an alternative freezing protocol for goat embryos of different morphology and quality was tested. Fifty-eight embryos on Day 6-7 stage were transferred as fresh or after freeze-thawing (n=29/group). For freezing, embryos were placed into 1.5M ethylene-glycol solution for 10min. During this time, they were loaded in the central part of 0.25mL straw, separated by air bubble from columns containing PBS/BSA 0.4% plus 20% BFS. Straws were then frozen using a freezing machine from 20ºC to -6ºC at a cooling rate of 3ºC/min, stabilization for 15min (seeding after 5min), from -6 C to -32ºC at 0.6 C/min,and plunged into liquid nitrogen. Frozen embryos were thawed for 30s at 37ºC in a water bath. Embryos subjected to fresh transfer were maintained in holding medium (37ºC). Fresh and frozen-thawed embryos were transferred at day 7 post-estrus to 30 recipients. Kidding and kid born rates were similar (P> 0.05), respectively, for recipients receiving fresh (66.7% or 10/15; 55.2% or 16/29) or frozen-thawed (60% or 9/15; 51.7% or 15/29) embryos. The cryopreservation of goat embryos using slow-freezing protocol and 1.5MEG resulted in similar efficiency rates of fresh embryos.

Combined treatment with oestradiol benzoate, d‐cloprostenol and oxytocin permits cervical dilation and nonsurgical embryo recovery in ewes

“…Earlier studies have shown that treatment of goats with PGF 2 α (Fonseca et al., ), PGF 2 α+OT (Pereira, Sohnrey, & Holtz, ) or ewes with EB+OT (Wulster‐Radcliffe, Costine, & Lewis, ) prior to embryo recovery did not adversely affect embryo quality. Transfer of embryos recovered from donor goats pretreated with cloprostenol for 16 hr prior to embryo recovery resulted in nearly 60% pregnancy rate when both the fresh or frozen‐thawed, good and excellent quality embryos were used (Fonseca et al., ). It will be interesting to evaluate the quality of the ovine embryos generated in this experiment with ensuing transfers or in vitro trials.…”

Pregnancy rate after fixed‐time transfer of cryopreserved embryos collected by non‐surgical route in Lacaune sheep

“…Embryo morphology was classified according to the IETS. Grade I and II embryos randomly underwent either slow freezing (SF; Fonseca et al, ) or vitrification (VT; Gibbons, Bruno‐Galarraga, Fernandez, Gonzalez‐Bulnes, & Cueto, ). The percentage of morulae and blastocysts transferred did not differ ( p < .05, chi‐square test) in SF (48.4%–30/62 and 51.6%–32/62) and VT (36.4%–16/44 and 63.6–28/44) groups, respectively.…”

“…Embryo transfer was performed using the semi‐laparoscopic technique on Day 8.5 to the horn ipsilateral to the CL. After 24 hr fasting, recipients that presented at least 1 Cl (92%; 58/63) received one or two cryopreserved embryos (morulae and/or blastocysts) (Fonseca et al, ). Pregnancy rate (number of pregnant recipients/number of recipients receiving embryos) and embryo survival rate (number of embryos with heartbeat/number of embryos transferred) were recorded by ultrasonography on Day 31.…”

Ultrasonographic cervical evaluation: A tool to select ewes for non‐surgical embryo recovery

“…. The hormonal combination of d-cloprostenol, estradiol benzoate and oxytocin was tested in Brazilian sheep breeds (Fonseca, et al, 2019;Fonseca et al, 2019aFonseca et al, , 2019b, achieving high rates of cervical transposing, uterine flushing and media recovered.…”

“…However, the feasibility of cervical access and manipulation (i.e. immobilization, retraction and penetrability) should be tested in individual donor ewes before any embryo collection attempt, aiming better effectiveness in the application of NSER (Fonseca, 2017;Fonseca et al, 2019aFonseca et al, , 2019bSantos et al, 2019).…”