Testing the efficacy and efficiency of a single “universal warming protocol” for vitrified human embryos: prospective randomized controlled trial and retrospective longitudinal cohort study

Abstract

PurposeTo study the efficacy and efficiency of a “universal warming protocol” for vitrified human embryos, based on subsequent steps with 1 and 0.5 M concentration of extracellular cryoprotectant (ECCP).MethodTwo studies on patients undergoing fertility treatments via ICSI: a prospective randomized controlled trial (RCT) and a retrospective cohort study (CS). Setting: Private assisted reproductive (AR) center.RCT: duration 01/03/2017–01/10/2017; 315 embryos at blastocyst stage obtained from 169 patients. Each patient’s embryos were first randomized for vitrification with two different kits: Vitrification Kit (Kitazato, Japan) and Sage Vitrification Kit (Origio, Denmark). The embryos were randomly warmed with either Kitazato (K) or Sage (S) warming kits, specifically: group A (KK), group B (KS), group C (SK), and group D (SS). Primary outcome measure: survival rate (number of embryos surviving per number of embryos warmed). Secondary: implantation rate (number of embryos implanted per number of embryos transferred).CS: duration 01/01/2013–31/12/2015 embryos from patients’ own oocytes; 10/04/2015–31/07/2017 embryos from donors’ oocytes. A total of 1055 embryos vitrified at cleavage stage obtained from 631 warming cycles: 847 of these obtained from patients’ own oocytes, 208 egg-donation-derived embryos. Each patient’s embryos were vitrified and warmed in various combinations of three different vitrification/warming kits: Kitazato (K), Sage (S), or made in-house in our laboratory (H). Vitrification/warming kits from different manufacturers are routinely used in our AR center, and the warming procedures are randomly performed with any available kit on a “first-in-first-out” basis, irrespective of the kit used for vitrification. Group names: KK, KS, SK, SS, SH, HK, HS, HH (embryos from patients’ own oocytes); eKK, eKS, eSK, eSS (egg-donation-derived embryos).ResultsCryo-survival rates were comparable in all study groups.RCT. Group A 99.0% (96/97), group B 98.8% (83/84), group C 98.4% (61/62), and group D 98.6% (71/72).CS. Embryos from patients’ own oocytes: KK 96.4% (54/56), KS 100.0% (13/13), SK 98.8% (80/81), SS 97.2% (174/179), SH 97.6% (40/41), HK 95.2% (20/21), HS 99.5% (187/188), and HH 97.4% (261/268). Egg-donation-derived embryos: eKK 100.0% (91/91), eKS 98.4% (60/61), eSK 100.0% (26/26), and eSS 96.7 (29/30).Implantation was generally comparable in all study groups—exceptions were in CS: KS vs. SK (P = 0.049), SS (P = 0.012), HS (P = 0.010), HH (P = 0.025); and SH vs. SS (P = 0.042), HS (P = 0.035).ConclusionWorldwide, millions of embryos have been cryopreserved using different vitrification kits; these studies establish that it is possible to combine different kits for vitrification and warming using a universal warming protocol. This can optimize costs, simplify lab routines, and favor embryo exchange between IVF centers.RCT registration numberISRCTN12342851.

‘Two countries-two labs’: the transnational gamete donation (TGD) programme to support egg donation

“…The medium was changed on day 3 when embryos were cultured to day 5 (ORIGIO® Sequential Blast TM). Supernumerary embryos were vitrified on day 3 or day 5, using the Cryotop method for embryo vitrification [ 13 , 14 ].…”

“…Multiple gestational sacs were counted as one CP. Implantation rate (IR) was defined as the number of gestational sacs divided by the number of transferred embryos; 'CP' as the presence of a gestational sac, with or without a fetal heartbeat, on ultrasonography; 'ongoing pregnancy' as a pregnancy beyond 12-week gestation; and 'live birth' as the delivery of one or more living infants [14,16].…”

A monocentric analysis of the efficacy of extracellular cryoprotectants in unfrozen solutions for cleavage stage embryos

“…In this perspective, kit 3 fits better both features of vitrification compared to kits 1 and 2, resulting in accordance with the rationale of a previous study demonstrating that, irrespective of the freezing protocol, a rapid warming procedure by stepwise dilutions of extracellular cryoprotectants only, as sucrose, may be adopted for higher survival rate of both slow-frozen and vitrified reproductive cells [34]. The use of only extracellular cryoprotectant (sucrose or threalose) in the thawing solution seems to be the best option for any frozen cell or tissue, as confirmed by the studies on clinical efficiency of Parmegiani’s Universal Warming [40, 41].…”

Effect of embryo cryopreservation duration on pregnancy-related complications and birthweight after frozen-thawed embryo transfer: a retrospective cohort study

“…Before November 9, 2015, the embryo vitrification and thawing kit of JieYing Laboratory Inc (Longueuil, Quebec, Canada) was applied, 18,19 and after that, the Cryotop® of Kitazato BioPharma Co. Ltd (Fuji, Japan) was used. 20 The operation procedures were in accordance with the manufacturer's instructions. When vitrification: 1) Transfer embryos to Equilibration Solution (ES) for 5 min (JieYing Kit)/ 7-8 min (Cryotop® Kit) at room temperature; 2) Transfer embryos to Vitrification Solution (VS) with minimal volume of ES and equilibrate for 1 mi (JieYing Kit)/30-60 s (Cryotop® Kit); 3) Quickly place the embryos on the JY straw/Cryotop straw with minimal volume of VS (each straw contains 1-2 embryos); 4) Plunge the straw into sterile liquid nitrogen and fit the straw cap; 5) Transfer the straw to storage dewar (MVE XC47/11-6SQ, Chart Industries, GA, USA).…”