Effects of cryoprotectant and plunging temperature in liquid nitrogen on the in vitro and in vivo development of murine morulae
The effects of plunging temperature in liquid nitrogen and cryoprotectant dilution methods were evaluated for compacted mouse morulae frozen in 1.5 M ethylene-glycol (E), 1.5M propylene-glycol (P) or 1.4M glycerol (G). Morulae were equilibrated for 10 minutes in cryoprotectant solution and loaded into 0.25 ml straws with cryoprotectant solution in 3 columns (groups E1, P1, G1) or cryoprotectant in the center and PBS in the lateral columns (E2, P2). Straws were cooled at 0.5ºC/min to -25 or -30ºC and plunged into liquid nitrogen. Straws were thawed in water at 22ºC for 20 seconds. Cryoprotectant was diluted in 3 steps for group G1 and in one step for groups E1 and P1 (direct transfer to PBS + 10% FCS) and E2 and P2 (shaken to mix the 3 columns before transferring to PBS+ 10% FCS). Plunging temperature had no significant effect on the proportion of morulae developed to blastocysts in vitro; this proportion was higher (p < 0.0001) in E1 (69.2%) than in E2 (60.3%), G1 (51.9%) and combined for P1 and P2 (46.9%). In second experiment, the proportion of transferred morulae that developed to viable fetuses was lower (p < 0.07) for E1-25 than for E1-30, G1-30, E2-25 or unfrozen (control) embryos (8.7, 20.0, 20.0, 17.4 and 19.8%, respectively). In conclusion, the ethylene glycol diluted directly in PBS (E1) exhibit the highest rate of in vitro embryos development, but based on in vivo embryos development was more efficacious in plunging temperature at -30ºC (E1-30).
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Faculdade de Medicina Veterinária e Zootecnia / Universidade de São Paulo
2000
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oai:scielo:S1413-959620000003000122001-02-07Effects of cryoprotectant and plunging temperature in liquid nitrogen on the in vitro and in vivo development of murine morulaePANTANO,ThaísMELLO,Marco Roberto Bourg deGARCIA,José FernandoHO,Linda LeeVISINTIN,José Antonio Embryos Cryopreservation Glycerol Ethylene glycol Propanediols The effects of plunging temperature in liquid nitrogen and cryoprotectant dilution methods were evaluated for compacted mouse morulae frozen in 1.5 M ethylene-glycol (E), 1.5M propylene-glycol (P) or 1.4M glycerol (G). Morulae were equilibrated for 10 minutes in cryoprotectant solution and loaded into 0.25 ml straws with cryoprotectant solution in 3 columns (groups E1, P1, G1) or cryoprotectant in the center and PBS in the lateral columns (E2, P2). Straws were cooled at 0.5ºC/min to -25 or -30ºC and plunged into liquid nitrogen. Straws were thawed in water at 22ºC for 20 seconds. Cryoprotectant was diluted in 3 steps for group G1 and in one step for groups E1 and P1 (direct transfer to PBS + 10% FCS) and E2 and P2 (shaken to mix the 3 columns before transferring to PBS+ 10% FCS). Plunging temperature had no significant effect on the proportion of morulae developed to blastocysts in vitro; this proportion was higher (p < 0.0001) in E1 (69.2%) than in E2 (60.3%), G1 (51.9%) and combined for P1 and P2 (46.9%). In second experiment, the proportion of transferred morulae that developed to viable fetuses was lower (p < 0.07) for E1-25 than for E1-30, G1-30, E2-25 or unfrozen (control) embryos (8.7, 20.0, 20.0, 17.4 and 19.8%, respectively). In conclusion, the ethylene glycol diluted directly in PBS (E1) exhibit the highest rate of in vitro embryos development, but based on in vivo embryos development was more efficacious in plunging temperature at -30ºC (E1-30).info:eu-repo/semantics/openAccessFaculdade de Medicina Veterinária e Zootecnia / Universidade de São PauloBrazilian Journal of Veterinary Research and Animal Science v.37 n.3 20002000-01-01info:eu-repo/semantics/articletext/htmlhttp://old.scielo.br/scielo.php?script=sci_arttext&pid=S1413-95962000000300012en10.1590/S1413-95962000000300012 |
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PANTANO,Thaís MELLO,Marco Roberto Bourg de GARCIA,José Fernando HO,Linda Lee VISINTIN,José Antonio |
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PANTANO,Thaís MELLO,Marco Roberto Bourg de GARCIA,José Fernando HO,Linda Lee VISINTIN,José Antonio Effects of cryoprotectant and plunging temperature in liquid nitrogen on the in vitro and in vivo development of murine morulae |
author_facet |
PANTANO,Thaís MELLO,Marco Roberto Bourg de GARCIA,José Fernando HO,Linda Lee VISINTIN,José Antonio |
author_sort |
PANTANO,Thaís |
title |
Effects of cryoprotectant and plunging temperature in liquid nitrogen on the in vitro and in vivo development of murine morulae |
title_short |
Effects of cryoprotectant and plunging temperature in liquid nitrogen on the in vitro and in vivo development of murine morulae |
title_full |
Effects of cryoprotectant and plunging temperature in liquid nitrogen on the in vitro and in vivo development of murine morulae |
title_fullStr |
Effects of cryoprotectant and plunging temperature in liquid nitrogen on the in vitro and in vivo development of murine morulae |
title_full_unstemmed |
Effects of cryoprotectant and plunging temperature in liquid nitrogen on the in vitro and in vivo development of murine morulae |
title_sort |
effects of cryoprotectant and plunging temperature in liquid nitrogen on the in vitro and in vivo development of murine morulae |
description |
The effects of plunging temperature in liquid nitrogen and cryoprotectant dilution methods were evaluated for compacted mouse morulae frozen in 1.5 M ethylene-glycol (E), 1.5M propylene-glycol (P) or 1.4M glycerol (G). Morulae were equilibrated for 10 minutes in cryoprotectant solution and loaded into 0.25 ml straws with cryoprotectant solution in 3 columns (groups E1, P1, G1) or cryoprotectant in the center and PBS in the lateral columns (E2, P2). Straws were cooled at 0.5ºC/min to -25 or -30ºC and plunged into liquid nitrogen. Straws were thawed in water at 22ºC for 20 seconds. Cryoprotectant was diluted in 3 steps for group G1 and in one step for groups E1 and P1 (direct transfer to PBS + 10% FCS) and E2 and P2 (shaken to mix the 3 columns before transferring to PBS+ 10% FCS). Plunging temperature had no significant effect on the proportion of morulae developed to blastocysts in vitro; this proportion was higher (p < 0.0001) in E1 (69.2%) than in E2 (60.3%), G1 (51.9%) and combined for P1 and P2 (46.9%). In second experiment, the proportion of transferred morulae that developed to viable fetuses was lower (p < 0.07) for E1-25 than for E1-30, G1-30, E2-25 or unfrozen (control) embryos (8.7, 20.0, 20.0, 17.4 and 19.8%, respectively). In conclusion, the ethylene glycol diluted directly in PBS (E1) exhibit the highest rate of in vitro embryos development, but based on in vivo embryos development was more efficacious in plunging temperature at -30ºC (E1-30). |
publisher |
Faculdade de Medicina Veterinária e Zootecnia / Universidade de São Paulo |
publishDate |
2000 |
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http://old.scielo.br/scielo.php?script=sci_arttext&pid=S1413-95962000000300012 |
work_keys_str_mv |
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