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In frame A, pre-labeled CBS embryo straw awaits open-ended (right straw) within the confines of its sterile packaging during vitrification set-up. The central straw is being reliably sealed in an automated Syms 1 sealer, with the completed weld seal seen on the straw to the left. Note that the tamperproof internalized labels can be different
Source publication
A novel, aseptic closed system vitrification (VTF) technique for the cryopreservation of embryos and oocytes has been developed and clinically validated in this study. It combines the practicality of embryo-containing sterile flexipettes stored safely and securely with 0.3ml CBS™ embryo straws possessing weld seals. The cooling and warming rates of...
Contexts in source publication
Context 1
... validation of cooling and warming rates (Phase 1) was determined using a DATAQ Instruments Model DI-1000-USB data logger (www.dataq.com) and an Omega 5SRTC-TT-T-30-36 fine (0.13 mm diameter) thermocouple. The thermocouple was threaded into the base of the flexipette filled with 3 ll of I.C.E. vit- rification solution (n = 4), and the flexipette was then inserted into a 0.3 ml CBS straw, which was then plunged into LN 2 for tempera- ture tracking at 0.2 s intervals. Similar measurements were also taken upon warming, but for warming, the extracted flexipette was immediately allowed to warm in ambient air (preliminary study, Fig. 1) or plunged into a 37 °C solution within a 58 mm petri dish (Figs. 2C and 3). All data points were plotted using SigmaPlot, and mean cooling and warming rates were calculated for each run between 0 and À125 °C and averaged over four independent ...
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... vitrification procedure has been described in detail previously [32,33] (Fig. 2). All handling procedures were performed at room temperature (20-24 °C) using a 3-step vitrification solution addition and 5-step dilution procedure, as described in detail elsewhere [32]. For vitrification, all oocytes and embryos (day 1 and 3) were placed into V1 and V2 solutions for 5 min and 2 min, respectively, and 5 min and 5 min for blastocysts. Following V2 equilibration, embryos and oocytes were pipetted into the final V3 solution for 1-1.5 min, respectively, and loaded into a 3 ll column in the flexipette. The tip was simply removed and wiped dry on sterile gauze, and then inserted completely into the inner lumen of the open-ended straw before final weld sealing and direct placement into LN 2 ...
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... to cooling, the outer CBS straws were heat sealed as described in Fig. 2A. For warming, the identification of a patient's colorized straw label (with the aseptic contents submerged in LN 2 ) was safely and effectively performed by grasping the lS-VTF straw with large surgical scissors (Fig. 2B) just below the internal plug. Once the ID was confirmed, the straw was quickly pulled up into ambient air and a ''cut-tilt-tap'' technique applied (refer to YouTube video, ''microSecure vitrification warming''). The straw was cut in a horizontal position while holding the non-labeled end and then tilted downward (45-60° angle) over a warm (37 °C) 0.5 M sucrose bath, tapped gently and the vitrifica- tion tip was allowed to free-fall directly into the 58 mm diameter petri dish for rapid warming (Fig. 2C). Within 10 s, the flexipette was removed from the bath and its contents expelled into a 1.0 M sucrose solution. The cryoprotectants were then eluted from the oocytes or embryos in a series of declining sucrose concentra- tions (e.g., 1.0 M to 0.5 M to 0.25 M to 0.1 M; proprietary solutions: T1-T4; 5 min/step) at room temperature (20-24 °C) before a 5 min isotonic equilibration in culture medium at 37 °C, as described elsewhere [32,33]. The routine thawing of slow frozen blastocysts has been described previously [15]. Pregnancy outcomes were evaluated by the determination of +b-hCG levels, clinical pregnancy, implantation and live birth rates. In Phase 3, the cryopreservation treatments were all compared to a fresh blastocyst embryo transfer group with rate (%) differences determined using Chi-square ...
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... to cooling, the outer CBS straws were heat sealed as described in Fig. 2A. For warming, the identification of a patient's colorized straw label (with the aseptic contents submerged in LN 2 ) was safely and effectively performed by grasping the lS-VTF straw with large surgical scissors (Fig. 2B) just below the internal plug. Once the ID was confirmed, the straw was quickly pulled up into ambient air and a ''cut-tilt-tap'' technique applied (refer to YouTube video, ''microSecure vitrification warming''). The straw was cut in a horizontal position while holding the non-labeled end and then tilted downward (45-60° angle) over a warm (37 °C) 0.5 M sucrose bath, tapped gently and the vitrifica- tion tip was allowed to free-fall directly into the 58 mm diameter petri dish for rapid warming (Fig. 2C). Within 10 s, the flexipette was removed from the bath and its contents expelled into a 1.0 M sucrose solution. The cryoprotectants were then eluted from the oocytes or embryos in a series of declining sucrose concentra- tions (e.g., 1.0 M to 0.5 M to 0.25 M to 0.1 M; proprietary solutions: T1-T4; 5 min/step) at room temperature (20-24 °C) before a 5 min isotonic equilibration in culture medium at 37 °C, as described elsewhere [32,33]. The routine thawing of slow frozen blastocysts has been described previously [15]. Pregnancy outcomes were evaluated by the determination of +b-hCG levels, clinical pregnancy, implantation and live birth rates. In Phase 3, the cryopreservation treatments were all compared to a fresh blastocyst embryo transfer group with rate (%) differences determined using Chi-square ...
Context 5
... to cooling, the outer CBS straws were heat sealed as described in Fig. 2A. For warming, the identification of a patient's colorized straw label (with the aseptic contents submerged in LN 2 ) was safely and effectively performed by grasping the lS-VTF straw with large surgical scissors (Fig. 2B) just below the internal plug. Once the ID was confirmed, the straw was quickly pulled up into ambient air and a ''cut-tilt-tap'' technique applied (refer to YouTube video, ''microSecure vitrification warming''). The straw was cut in a horizontal position while holding the non-labeled end and then tilted downward (45-60° angle) over a warm (37 °C) 0.5 M sucrose bath, tapped gently and the vitrifica- tion tip was allowed to free-fall directly into the 58 mm diameter petri dish for rapid warming (Fig. 2C). Within 10 s, the flexipette was removed from the bath and its contents expelled into a 1.0 M sucrose solution. The cryoprotectants were then eluted from the oocytes or embryos in a series of declining sucrose concentra- tions (e.g., 1.0 M to 0.5 M to 0.25 M to 0.1 M; proprietary solutions: T1-T4; 5 min/step) at room temperature (20-24 °C) before a 5 min isotonic equilibration in culture medium at 37 °C, as described elsewhere [32,33]. The routine thawing of slow frozen blastocysts has been described previously [15]. Pregnancy outcomes were evaluated by the determination of +b-hCG levels, clinical pregnancy, implantation and live birth rates. In Phase 3, the cryopreservation treatments were all compared to a fresh blastocyst embryo transfer group with rate (%) differences determined using Chi-square ...
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Citations
... 9 With the popularization of vitrification and freezing technology, vitrification and freezing carriers have been developed and applied, further improving the effect of vitrification and freezing. This method uses many types of commercial [10][11][12] and non-commercial [13][14][15] vitrification carriers. Regardless of whether the embryos are directly in contact with liquid nitrogen when the embryos are stored in a liquid nitrogen tank after freezing, the carriers can be divided into three categories: fully open carriers, 16 open cooling and closed storage carriers, 17 and closed carriers. ...
Objective
This study aimed to compare the effects of two brands of commercial vitrification carriers on pregnancy outcomes in freeze–thaw cycles.
Methods
We included 4871 patients who underwent a “freeze all” strategy using the commercial carriers J.Y. straw and OYASHIPS straw in the Reproductive Center of the First Hospital of Jilin University. The pregnancy outcomes of cleavage-stage embryos and blastocysts were studied separately. Detailed data and the safety of children born from mothers with the two types of carriers were also compared.
Results
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Conclusion
The OYASHIPS straw carrier is cheap and can achieve clinical pregnancy and live birth outcomes comparable to those of J.Y. straw. Therefore, OYASHIPS straw is a good alternative option.
... This was primarily mediated by a misconception that ultra-rapid cooling of oocytes achieved by the direct plunging of oocytes and embryos into liquid nitrogen before protective capping (not sealed closed) was absolutely necessary to optimize a vitrified state that insured post-warming survival/viability 26 . It has since been shown in prospective, randomized studies that different closed system devices (e.g., HSV, Vitrisafe) can indeed achieve comparable oocyte survival and developmental competence rate to that of an open system [27][28][29] , and high blastocyst survival/live birth rates [30][31] , while maintaining complete security from vertical disease transmission 32 . ...
... Although the risk of viral transfection is unproven, risk assessment potential is virtually eliminated in an "aseptic, closed vitrification system" like the high security vitrification (HSV) 28,33 , microSecure vitrification (µS-VTF) [30][31] and Vitrisafe 27,29,32 approaches. Unlike other original sub-optimal closed designs approved by FDA, the latter systems are superior in that their carrier devices are inserted into CryoBioSystem (CBS) straws. ...
The global COVID-19 pandemic of 2020 to date has altered all our lives and how clinical medicine is practiced. Faced with a highly infectious coronavirus, specifically known as Severe Acute Respiratory Coronavirus-2 (SARS-CoV-2), in our homes, clinical environment and laboratories, we have applied preventative measures to potentially control and contain this mysterious pathogen. Although the concepts of frequent handwashing/ disinfection, social distancing and face mask wearing were rigidly applied with regional variation of acceptance in the USA, clinics and reproductive biology/IVF laboratories adapted strict global policies for a safer workplace for our staff, patients, and the specimens we handled. We address the concerns that resurfaced with the COVID -19 pandemic, regarding potential disease transmission between hosts, reproductive tissue (sperm, oocytes and embryos), recipient uteri and the fetus. To what extent were preventative measures sufficient and is there a need for adopting “best practices” above and beyond established “good tissue practices”. In fear of future pandemic disease events (“pandemic-mania”) impacting fertility treatments, this paper addresses the rationale and benefits of adhering strictly to best practices, like the use of secure closed system biocontainment in cryostorage as an important preventative measure. Additionally, historical and current perspectives are discussed and the ability to change established practices under the guise of commercial influences.
... Among these technologies, in vitro embryo production (IVEP) is an important technique that has great potential for genetic enhancement [1] through conservation of high-quality germline and transferring them over generations. During oocyte maturation and embryo culture, various factors are crucial to oocyte developmental competence, embryo quality, and cryotolerance, such as lipid content, lipid composition, embryo metabolism, apoptosis, gene expression pattern [2], optimized culture media composition (additives, salts, amino acids, hormones, sugars, antioxidants, pH, and osmolarity), temperature, oxygen tension, and oocyte donor and semen quality [3]. As an integral event during metabolism, oocytes use the energy derived from lipid droplets located in the cumulus-oocyte complex through the beta-oxidation pathway for subsequent metabolic processes [4]. ...
Background and Aim: Despite many trials, buffalo embryos have poor cryosurvivability because of their high lipid content. L-carnitine was found to be a lipid-reducing agent when added to oocyte and embryo culture media. The study aimed to determine the most effective concentration of L-carnitine to improve the oocyte developmental competence and cryotolerance of buffalo embryos.
Materials and Methods: In vitro maturation and embryo culture media were supplemented with four concentrations of L-carnitine: 0 (control), 0.25, 0.5, and 1 mM. Good-quality embryos on 7 days were vitrified using mixtures of dimethyl sulfoxide and ethylene glycol at two concentrations (3.5 and 7 M).
Results: The result showed that the cleavage and morula rates were significantly (p
... The cooling and warming rates in the GAVI pod estimated at 14,100 °C/min and 11,000 °C/min, respectively [14] which are significantly higher than those reported for closed vitrification systems with high security CBS straws (1000 to 2000 °C/min) [20]. Those cooling rates were also validated in another closed CBS straw system [21]. Therefore, the potential benefit of the higher cooling rate with closed system GAVI pods compared to conventional closed-type systems such as HSV CBS on embryo viability is considered small, and as a result may not have affected clinical outcomes in this study. ...
PurposeDoes semi-automated vitrification have lower inter-operator variability and better clinical outcomes than manual vitrification?Methods
Retrospective analyses of 282 patients whose embryos had been cryopreserved, manually with Irvine®-CBS® (MV) or semi-automatically vitrified with the GAVI® method (AV) (from November 2017 to September 2020). Both techniques were performed during the same period by 5 operators. Inter-operator variability was statistically analyzed between operators who performed the vitrification and those who performed the warming process to compare the intact survival rate (% embryos with 100% intact blastomeres) and the positive survival rate (at least 50% intact blastomeres). Additionally, the complete vitrification time was assessed for the 2 techniques according to the number of vitrified embryos.ResultsManual vitrification involved warming 338 embryos in 266 cycles for 181 couples compared to 212 embryos in 162 AV cycles for 101 patients. The positive survival rate was higher (p < 0.05) after MV (96%; 323/338) than after AV (90%; 191/212). The intact survival rate (86 vs 84%) and the clinical pregnancy rate (27 vs 22%) were not significantly different between MV and AV. Regarding the inter-operator variability, no significant difference in positive and intact survival rate was evident between the 5 technicians, neither by vitrification nor by warming steps with MV and AV. Concerning time-saving, the MV technique proved to be quicker than AV (minus 11 ± 9 min).Conclusions
Manual vitrification exhibited favorable total survival rates and was more time efficient, while both MV and AV cooling and warming treatments showed little operator variability.
... Furthermore, in the literature, there are reported vitrification systems that do not require high cooling/heating rates and small volumes to obtain vitrified embryos or oocytes (such as I.C.E. Vitrification, Innovative Cryo Enterprises LLC, Linden, NJ) [39][40][41]. Despite the successful use of "open" carriers for vitrification, direct contact between the sample and LN 2 may not be desirable because of the potential risk of cross-contamination between specimens and inadvertent exposure to contaminants present in the tanks [42][43][44]. ...
Purpose
The main purpose and research question of the study are to compare the efficacy of high-security closed versus open devices for human oocytes’ vitrification.
Methods
A prospective randomized study was conducted. A total of 737 patients attending the Infertility and IVF Unit at S.Orsola University Hospital (Italy) between October 2015 and April 2020 were randomly assigned to two groups. A total of 368 patients were assigned to group 1 (High-Security Vitrification™ - HSV) and 369 to group 2 (Cryotop® open system). Oocyte survival, fertilization, cleavage, pregnancy, implantation, and miscarriage rate were compared between the two groups.
Results
No statistically significant differences were observed on survival rate (70.3% vs. 73.3%), fertilization rate (70.8% vs. 74.9%), cleavage rate (90.6% vs. 90.3%), pregnancy/transfer ratio (32.0% vs. 31.8%), implantation rate (19.7% vs. 19.9%), nor miscarriage rates (22.1% vs. 21.5%) between the two groups. Women’s mean age in group 1 (36.18 ± 3.92) and group 2 (35.88 ± 3.88) was not significantly different (P = .297). A total of 4029 oocytes were vitrified (1980 and 2049 in groups 1 and 2 respectively). A total of 2564 were warmed (1469 and 1095 in groups 1 and 2 respectively). A total of 1386 morphologically eligible oocytes were inseminated by intracytoplasmic sperm injection (792 and 594 respectively, P = .304).
Conclusions
The present study shows that the replacement of the open vitrification system by a closed one has no impact on in vitro and in vivo survival, development, pregnancy and implantation rate. Furthermore, to ensure safety, especially during the current COVID-19 pandemic, the use of the closed device eliminates the potential samples’ contamination during vitrification and storage.
... Although the risk of viral transfection is unproven, risk assessment potential is virtually eliminated in an "aseptic, closed vitrification system" like the high security vitrification (HSV) [27], microSecure vitrification (μS-VTF) [28] [29], and Vitrisafe [30] [31] [32] approaches. Unlike other original sub-optimal closed designs approved by FDA, the latter systems are superior in that their carrier devices are inserted into CryoBioSystem's (CBS) straws. ...
Examine good tissue practices as relates to in vitro fertilization, biopsying, and vitrificationto compare current knowledge of ova, sperm, and embryos as vectors for disease transmission as it relates to our current knowledge regarding the SARS-CoV-2 virus.
Unknown risks relating to the SARS-CoV-2 virus and sperm, ova, and embryos necessitate a reexamining of how human IVF is performed. Over the last decade, improvements in cryosurvival and live birth outcomes have been associated with zona pellucida breaching procedures (e.g., blastocyst collapsing and biopsying). In turn, today embryos are generally no longer protected by an intact zona pellucida when vitrified and in cryostorage. Additionally, high security storage containers have proven to be resilient to potential cross-contamination and reliable for routine human sperm freezing and embryo vitrification.
Several options to current IVF practices are presented that can effectively mitigate the risks of cross-contamination and infection due to the current Covid-19 pandemic or other viral exposures. The question remains; is heightened security and change warranted where the risks of disease transmission likely remain negligible?
... In order to overcome the above scenarios, several solutions may be applicable: (1) The use of closed system vitrification systems incorporating CBS straws or other systems [5]. (2) The use of sterile LN and vapor storage [6]. ...
... Blastocyst formation was documented and typically only those possessing ≥2BB grade were cryopreserved or may have first underwent TE biopsy procedures for PGT-A determination if ≥3BB grade, as previously detailed [41,43]. All fair to excellent quality blastocysts were vitrified using our microSecure vitrification (μS-VTF) procedure [44,45]. Embryos were transferred on Day 3 or 5 depending on the number/quality of embryos available and the IVF history of the patient. ...
... 4), we were even more confident to strive toward a practice of single embryo transfer as advocated by Gardner and Lane [52] years earlier. Certainly, the concurrent development, validation, and implementation of highly reliable vitrification [44,45] and blastocyst biopsy [41,43] procedures has allowed us to confidently implement single ET with unprecedented success, as reflected in the outcome verification of our FET cycles in Figure 2B. Note that implantation success and live birth rates are similar within age groups and each occurs at an equally high level across age groups when transferring a mean of 1 to 1.2 blastocysts/FET. ...
... Based on our interests to optimize our culture media, we identified and added a highly purified recombinant, high-molecular weight HA product (EAS) which we felt had significant potential merits as a serumfree supplement to LG (mLG). Our studies validate that this EAS was indeed not harmful and is more likely beneficial based on the outstanding implantation and live birth rates we achieved, which has attained elite levels (i.e., upper 5 percentile) as previously demonstrated by our lab [41,44,45]. In addition to its published role as an EAS which enhances implantation [60], we believe strongly that HA promotes membrane plasticity, aiding in cellular healing and protective functions associated with stressful events like trophectoderm biopsy and blastocyst vitrification. ...
Objective: To develop and validate a reliable in vitro culture system for human embryos. Design: Retrospective analyses of a series of four studies were conducted between 2006 and 2010 to assess the effect of incubator type (CO2 box versus Tri-gas minibox), media type, oil type, and hyaluronate supplementation. Optimization of in vitro blastocyst development was verified by assessing our National CDC/ART Surveillance reports between 2010 and 2016. Material and Methods: All patients experienced controlled ovarian hyperstimulation, followed by egg retrieval 35 h post-hCG. Cumulus-oocyte complexes were temporarily cultured in P1 or LG Fert medium plus HSA. Eggs were moved to a more complex media (G-medium or Global®-LG medium) containing a synthetic protein and embryo adhesion supplement (SPS and EAS, respectively; mLG) post-ICSI insemination. Zygotes were assigned to group culture in 25 µl droplets under oil (light mineral oil or paraffin oil; 37 °C) and embryo development was evaluated on Days 3, 5, and 6 and transferred on Day 3 to 5 depending on the number/quality of embryos available and the IVF history of the patient. Transfers were performed under ultrasound guidance, primarily using a Sureview-Wallace catheter, and enriched ET medium containing 500 µg/mL EAS. Results: Pilot study results (Expt. 1) showed that a mLG single-step medium could be effectively used in combination with Sanyo MCO-5 tri-gas (TG) incubators. Once adapted to SCIRS Lab in 2007 (Expt. 2), the latter culture system yielded improved blastocyst production and pregnancy outcomes compared to CO2 in air sequential incubation in P1/Multi-blast medium. In Expt. 3, the mLG/TG system yielded high levels of ≥2BB quality blastocysts (51 to 66%) across all age groups, and greater (p < 0.05) pregnancy success/live birth rates using fewer embryos transferred on Day 5 versus Day 3. After validating its clinical effectiveness, mLG was then prospectively compared to a new generation G-media (1.5 & 2.5; Expt. 4) and determined that the crossover treatment using paraffin oil (Ovoil™) allowed the mLG system to be optimized. Subsequently, a compilation of our Annual CDC/ART reported data over six years verified the overall viability of in vitro cultured and vitrified blastocysts produced in the mLG/TG system. Conclusion: By systematically evaluating and implementing various components of an embryo culture system we were able to optimize blastocyst development over the last decade. Our mLG/TG culture system modified an exceptionally well designed KSOMAA LG medium using endotoxin-free EAS and SPS additives to support cellular membrane wellness under stressful in vitro conditions (e.g., culture, cell biopsy, vitrification). Our use of the mLG/TG culture system has proven to be effective, creating reliably high blastocyst production, implantation, and healthy live births.
... [55] Only one study has verified the clinical efficacy of a noncommercial, aseptic closed vitrification system called microSecure for human embryos. [56] It is a low-cost, non-commercial, aseptic, closed system that offers technical simplicity and is reliable with excellent embryo survival and sustained viability. [56] More RCTs are required to show similar efficiency of closed systems and open carriers for cryopreservation. ...
... [56] It is a low-cost, non-commercial, aseptic, closed system that offers technical simplicity and is reliable with excellent embryo survival and sustained viability. [56] More RCTs are required to show similar efficiency of closed systems and open carriers for cryopreservation. Probably, in future storage of cells in the vapor phase of nitrogen instead of liquid nitrogen can be an option. ...
... SUPPORT OBJECTIVE: MicroSecure vitrification (mS-VTF) evolved as a non-commercial, FDA compliant device integrating sterile flexipettes (300mmID; Cook Medical) into CBSÔ embryo straws. The mean cooling rate (1391 C/min) and warming rate (6233 C/min) of this closed, aseptic VTF system were verified by Dr. Greg Fahy (Schiewe et al., 2015). In 2014, CBS removed traditional hydrophobic plugged embryo straws from the worldwide marketplace, being replaced by 0.3ml semen straws. ...
