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Ashok Agarwal - One of the best experts on this subject based on the ideXlab platform.
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p 183 a comparative study on test yolk buffer and human Sperm Preservation medium on post thaw characteristics of human Sperm from pre freeze specimens
Fertility and Sterility, 2006Co-Authors: J Bhattacharya, S Kundu, Alex C Varghese, S M Bhattacharya, Asok K Bhattacharyya, Ashok AgarwalAbstract:J. Bhattacharya, S. Kundu, A. C. Varghese, S. M. Bhattacharya, A. K. Bhattacharyya, A. Agarwal; IVF & Infertility Research Center, Calcutta, India, University of Calcutta, Calcutta, India, Vivekananda Institute of Medical Sciences, Calcutta, India, Cleveland Clinic, Cleveland, OH Objective: Optimum cryosurvival is dependent not only on freezing methodologies, but also on the type of cryopreservatives used. It seems reasonable to speculate that removing the seminal plasma and selecting highly motile Spermatozoa by an optimal Sperm processing technique, before freezing, may reduce the loss of viability, motility and acrosome integrity during cryoPreservation. Moreover, this method can remove those cells that produce free radicals thereby minimizing peroxidative damage during freeze-thaw procedure. It has been shown that pre-freezing of Sperm before their preparation does not impair the ability of thawed Spermatozoa binding to zona pellucida. The objective of the present study was to compare the efficacy of TEST-Yolk buffer (TYB) and Human Sperm Preservation Medium (HSPM) during freeze-thaw process on pre-freeze prepared human Sperm. Design: Prospective study. Materials and Methods: Semen samples (n = 22) were obtained from healthy volunteers and preliminary analysis were done after liquefaction using Makler Sperm counting chamber and morphology using Papanicolaou staining. Samples were prepared using density gradient method and the final Sperm pellet suspended in Quinn’s Sperm washing media and divided into two aliquots. The aliquots were frozen with TYB medium (Irvine Scientific, Santa Ana, CA) and HSPM medium (Sperm Freeze™, FertiPro, Belgium) respectively and frozen in liquid nitrogen. Post thaw motility characteristics and morphology were analyzed both after thawing and following removal of cryoprotectants by density gradient preparation. Results: Though there were significant changes in various motility characteristics and morphology between prefreeze samples and samples frozen with either TYB or HSPM as given in table 1, there were no statistically significant changes in the post-thaw Sperm between the two groups. Moreover the Sperm characteristics following post thaw preparation showed no significant changes between the two groups (table 2). Conclusion: TYB and HSPM mediums are effective in cryoPreservation protocols when pre-freeze semen samples are used for Sperm preparation. Although there was a lack of statistically significant difference between the two groups, the post thaw motility patterns showed better values in samples frozen in TYB medium. Financial Support: None. Print this Page for Your Records Close WindowTable 1: Changes in pre-freeze and post-thaw semen characteristics after freezing with TYB and HSPMOasis, The Online Abstract Submission System Page 1 of 3http://www.abstractsonline.com/submit/SubmitPrinterFriendlyVersion.asp?ControlKey=%7... 5/1/2006
W Schmidt - One of the best experts on this subject based on the ideXlab platform.
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comparison between human Sperm Preservation medium and test yolk buffer on protecting chromatin and morphology integrity of human Spermatozoa in fertile and subfertile men after freeze thawing procedure
Journal of Andrology, 2001Co-Authors: M E Hammadeh, S Greiner, P Rosenbaum, W SchmidtAbstract:: The aim of this study was to identify the detrimental effect of the freeze-thaw process on chromatin integrity and morphology of human Spermatozoa, and to determine whether human Sperm Preservation medium (HSPM) or TEST-yolk buffer (TYB) offers a better protection to Spermatozoa from cryodamage after the freeze-thaw procedure. Thirty-five semen samples obtained from couples childless because of male factor infertility (subfertile men, group 1) and 25 semen samples from healthy, normal volunteers of proven fertility (group 2) were included in the study. Each semen sample was divided into 2 parts, the first part was mixed with HSPM and the other with TYB (1:1), and frozen with a controlled slow-stage freezer, before plunging into liquid nitrogen. Twelve smears from each semen sample were made before (n = 4) and after (n = 8) the freeze-thaw process. Chromatin structure was evaluated after staining using the acridine orange (AO) test, whereas morphology was analyzed according to strict criteria. The mean percentage of Spermatozoa that exhibited normal morphology and intact chromatin structure was decreased after freeze-thaw in all samples treated with HSPM or TYB in comparison with the value observed in the native semen samples of both groups. However, TYB preserved chromatin and morphology significantly better than HSPM did (9.3% +/- 5.6% and 88.7% +/- 11.2% vs. 7.8% +/- 4.2% and 85.5% +/- 12.5%, respectively). Therefore, TYB could be recommended as a first choice cryoprotectant for semen Preservation in order to avoid extra chromatin structure damage and morphology alterations of Spermatozoa not only for patients pursuing assisted reproduction, but also for donor samples.
Takehito Kaneko - One of the best experts on this subject based on the ideXlab platform.
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Development of feline embryos produced using freeze-dried Sperm.
Theriogenology, 2020Co-Authors: Yasunori Tsujimoto, Takehito Kaneko, Takumi Yoshida, Kazuto Kimura, Toshio Inaba, Kikuya Sugiura, Shingo HatoyaAbstract:Abstract Freeze drying has been developed as a new Sperm Preservation method that eliminates the necessity of using liquid nitrogen. An advantage of freeze-dried Sperm is that it can be stored at 4 °C and transported at room temperature. To develop assisted reproductive techniques (ARTs) for domestic cats, we evaluated the effect of the freeze-dry procedure on cat Sperm DNA by analyzing DNA integrity (experiment 1) and by generating cat embryos using freeze-dried Sperm that had been preserved for several months (experiment 2). In experiment 1, the rate of DNA damage to freeze-dried Sperm was not significantly different than that of Sperm cryopreserved with liquid nitrogen (P > 0.05). In experiment 2, the proportions of cleaved embryos, morulae, and blastocysts and the cell number of blastocysts did not differ between experimental groups in which fresh Sperm and freeze-dried Sperm were used (P > 0.05). In addition, we generated feline blastocysts using freeze-dried Sperm stored for 1–5 months. These results support an expansion of the repertoire of ARTs that are potentially applicable to both domestic and endangered species of cats.
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322 Sperm Preservation by freeze drying in endangered animals
Reproduction Fertility and Development, 2015Co-Authors: Takehito KanekoAbstract:Sperm Preservation is a useful tool for conservation of endangered animals. Freeze-drying Sperm have been studied as new Preservation method in various mammals as samples can be preserved in a refrigerator at 4°C or ambient temperature. Sperm Preservation by freeze-drying is the ultimate method by which Sperm can be stored that neither required specialised cryoprotectants nor constant supply of liquid nitrogen. We established the freeze-drying method that mouse and rat Sperm could be preserved long-term at 4°C after freeze-drying using a simple solution containing 10 mM Tris and 1 mM EDTA (TE buffer; 2012 PLoS ONE 7, e35043; 2012 Cryobiology 64, 211–214). Using this method, the fertility of the chimpanzee, giraffe, and jaguar Sperm after freeze-drying were estimated. Ejaculated chimpanzee and giraffe and cauda epididymal jaguar Sperm were freeze-dried using TE buffer. Sperm were rehydrated with sterile distilled water after storage at 4°C for 1 month. Sperm with normal shape were injected into mouse oocytes in CZB medium with HEPES, and oocytes were then cultured in vitro for 6 to 8 h in the same media. In all animals, pronuclei and Sperm tail were observed into oocytes without artificial activation after injection of freeze-dried Sperm. When chimpanzee, giraffe, and jaguar Sperm were injected into oocytes, 86% (12/14), 100% (12/12), and 96% (22/23) of oocytes formed 2 distinct pronuclei. This study demonstrated that the Sperm of various animals could be decondensed into the mouse oocytes after freeze-drying using the same protocol. A further advantage is that freeze-dried Sperm can be transported oversea at ambient temperature. Freeze-drying Preservation without using liquid nitrogen can be protected strongly valuable gametes of endangered animals even in the event of unexpected accidents and disaster such as earthquakes and typhoons. Freeze-drying of Sperm has been applied as a “freeze-drying zoo” for conservation of endangered animals (http://www.anim.med.kyoto-u.ac.jp/reproduction/home.aspx).
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simple Sperm Preservation by freeze drying for conserving animal strains
Methods of Molecular Biology, 2015Co-Authors: Takehito KanekoAbstract:: Freeze-drying Spermatozoa is the ultimate method for the maintenance of animal strains, in that the gametes can be preserved for a long time in a refrigerator at 4 °C. Furthermore, it is possible to realize easy and safe transportation of Spermatozoa at an ambient temperature that requires neither liquid nitrogen nor dry ice. Freeze-drying Spermatozoa has been established as a new method for storing genetic resources instead of cryoPreservation using liquid nitrogen. This chapter introduces our latest protocols for freeze-drying of mouse and rat Spermatozoa, and the anticipated results of the fertilizing ability of these gametes following long-term Preservation or transportation.
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Sperm Preservation by freeze drying for the conservation of wild animals
PLOS ONE, 2014Co-Authors: Takehito Kaneko, Hidefusa Sakamoto, Manabu Onuma, Miho InouemurayamaAbstract:Sperm Preservation is a useful technique for the maintenance of biological resources in experimental and domestic animals, and in wild animals. A new Preservation method has been developed that enables Sperm to be stored for a long time in a refrigerator at 4°C. Sperm are freeze-dried in a solution containing 10 mM Tris and 1 mM EDTA. Using this method, liquid nitrogen is not required for the storage and transportation of Sperm. We demonstrate that chimpanzee, giraffe, jaguar, weasel and the long-haired rat Sperm remain viable after freeze-drying. In all species, pronuclei were formed after the injection of freeze-dried Sperm into the mouse oocytes. Although preliminary, these results may be useful for the future establishment of “freeze-drying zoo” to conserve wild animals.
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the latest improvements in the mouse Sperm Preservation
Methods of Molecular Biology, 2014Co-Authors: Takehito KanekoAbstract:: Sperm Preservation is an important technique for maintaining valuable genetic resources in biomedical research and wildlife. In the mouse, the Sperm cryoPreservation method has been established and adopted by large-scale Sperm Preservation projects in cryobanks. Recently, a new Sperm Preservation method using freeze-drying has been studied in various mammals. Freeze-drying is the ultimate method by which Sperm can be preserved long term in a refrigerator (4 °C). And it is possible to realize easy and safe transportation of Sperm at an ambient temperature that requires neither liquid nitrogen nor dry ice. Furthermore, it has been demonstrated that the fertilizing ability of Sperm cryopreserved or freeze-dried by the methods described in this chapter is well maintained during long-term Preservation. This chapter introduces the latest protocols for cryoPreservation and freeze-drying of mouse Sperm, and the anticipated results of the fertilizing ability of these Sperm preserved long-term.
D P Froman - One of the best experts on this subject based on the ideXlab platform.
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premises for fowl Sperm Preservation based on applied bioenergetics
Journal of Animal Science, 2014Co-Authors: D P FromanAbstract:: The primary goal of this work was to test whether the Sperm mobility assay could be used to derive mathematical relationships from which predictions could be made about Sperm cell function. A precondition was random sampling from a pool of Sperm. This precondition was met by centrifuging mobile Sperm through 12% (wt/vol) Accudenz containing the Ca(2+) chelator 1,2-bis-(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA) and then holding washed Sperm at 20°C within buffered potassium chloride. These 2 conditions rendered washed Sperm immobile at 20°C. Resumption of Sperm mobility was independent of time (P > 0.8558) when Sperm were reactivated at body temperature with 2 mM Ca(2+) in isotonic sodium chloride at pH 7.4. Reactivated Sperm mobility was 93% of the prewash control. Subsequent experiments served to define a dose response, predict optimal conditions for in vitro Sperm mobility, and show how Sperm can recover from an imposed non-physiological condition. Thus, functions were derived from which predictions were made. Whereas the utility of BAPTA treatment was confirmed in a new context, such utility did not address the question of whole-cell Ca(2+) flux during Sperm cell manipulation. This issue is pivotal for the application of bioenergetics to fowl Sperm Preservation. Therefore, the secondary goal of this research was to investigate Sperm cell Ca(2+) flux using a simulation of conditions encountered by Sperm during centrifugation through 12% (wt/vol) Accudenz. These conditions included a temperature of 30°C, a Ca(2+) sink, and no exogenous substrate. Sperm motion was measured with a Hobson SpermTracker. Data points conformed to parabolic functions when motile concentration and velocity were plotted as functions of time. In each case, maximums were observed, e.g., 26 min for motile concentration. The upswing was attributed to a redistribution of intracellular Ca(2+) whereas the downswing was attributed to Sperm cell Ca(2+) depletion. A pronounced isothermal increase was observed for each variable when the Ca(2+) sink was overcome with exogenous Ca(2+). Experimental outcomes supported four testable premises applicable to fowl Sperm Preservation research: 1) the importance of Sperm mobility phenotype, 2) the relationship between mitochondrial Ca(2+) cycling and Sperm mobility, 3) the utility of the Sperm mobility assay for predicting experimental outcomes, and 4) understanding mitochondrial Ca(2+) cycling in terms of whole-cell Ca(2+) flux.
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a new approach to Sperm Preservation based on bioenergetic theory
Journal of Animal Science, 2010Co-Authors: D P Froman, A J FeltmannAbstract:: To date, attempts to preserve chicken Sperm have been based on a trial-and-error experimental approach. The present work outlines the development of an alternative approach based on empiricism and bioenergetic theory. In previous work, we found fowl Sperm motility to be dependent on mitochondrial calcium cycling, phospholipase A(2), and long-chain fatty acids as an endogenous energy source. It is noteworthy that fowl Sperm reside within the Sperm storage tubules (SST) of the oviduct over an interval of days to weeks after insemination. In this regard, a model for in vivo Sperm storage was developed and tested in additional previous research. Sperm penetration of the SST, Sperm residence within the SST, and Sperm egress from the SST can be explained in terms mitochondrial function. Understanding Sperm function and longevity in terms of bioenergetics presented the possibility that Sperm could be inactivated by disrupting mitochondrial calcium cycling and could thereby be preserved. However, this possibility also posed a problem: maintenance of the inner membrane potential of the mitochondrion within inactivated Sperm. This report describes a series of experiments in which fowl Sperm were inactivated by treatment with the calcium chelator tetrasodium 1,2-bis-(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid, and then reactivated by treatment with calcium ions. The effect of tetrasodium 1,2-bis-(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid on mitochondrial calcium cycling was confirmed by flow cytometry and confocal microscopy. When treated Sperm were cooled to 10 degrees C, inactivated Sperm could be reactivated throughout a 5-h storage interval. When stored Sperm were held for 3 h before reactivation and insemination, fertility was 88% of the control. Storage did not affect hatchability. In summary, short-term storage was realized by manipulating mitochondrial function. We propose that 1) complex V consumes ATP within inactivated Sperm and, by doing so, maintains the inner membrane potential of the mitochondrion, 2) ATP is regenerated within inactivated Sperm by the action of creatine kinase on phosphocreatine, and 3) necrosis follows depletion of intracellular phosphocreatine. Therefore, future attempts to preserve chicken Sperm can be based on a theory that encompasses regulation of energy production, a biological context in which Sperm cells are motile, and the consequences of mitochondrial failure.
J Bhattacharya - One of the best experts on this subject based on the ideXlab platform.
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p 183 a comparative study on test yolk buffer and human Sperm Preservation medium on post thaw characteristics of human Sperm from pre freeze specimens
Fertility and Sterility, 2006Co-Authors: J Bhattacharya, S Kundu, Alex C Varghese, S M Bhattacharya, Asok K Bhattacharyya, Ashok AgarwalAbstract:J. Bhattacharya, S. Kundu, A. C. Varghese, S. M. Bhattacharya, A. K. Bhattacharyya, A. Agarwal; IVF & Infertility Research Center, Calcutta, India, University of Calcutta, Calcutta, India, Vivekananda Institute of Medical Sciences, Calcutta, India, Cleveland Clinic, Cleveland, OH Objective: Optimum cryosurvival is dependent not only on freezing methodologies, but also on the type of cryopreservatives used. It seems reasonable to speculate that removing the seminal plasma and selecting highly motile Spermatozoa by an optimal Sperm processing technique, before freezing, may reduce the loss of viability, motility and acrosome integrity during cryoPreservation. Moreover, this method can remove those cells that produce free radicals thereby minimizing peroxidative damage during freeze-thaw procedure. It has been shown that pre-freezing of Sperm before their preparation does not impair the ability of thawed Spermatozoa binding to zona pellucida. The objective of the present study was to compare the efficacy of TEST-Yolk buffer (TYB) and Human Sperm Preservation Medium (HSPM) during freeze-thaw process on pre-freeze prepared human Sperm. Design: Prospective study. Materials and Methods: Semen samples (n = 22) were obtained from healthy volunteers and preliminary analysis were done after liquefaction using Makler Sperm counting chamber and morphology using Papanicolaou staining. Samples were prepared using density gradient method and the final Sperm pellet suspended in Quinn’s Sperm washing media and divided into two aliquots. The aliquots were frozen with TYB medium (Irvine Scientific, Santa Ana, CA) and HSPM medium (Sperm Freeze™, FertiPro, Belgium) respectively and frozen in liquid nitrogen. Post thaw motility characteristics and morphology were analyzed both after thawing and following removal of cryoprotectants by density gradient preparation. Results: Though there were significant changes in various motility characteristics and morphology between prefreeze samples and samples frozen with either TYB or HSPM as given in table 1, there were no statistically significant changes in the post-thaw Sperm between the two groups. Moreover the Sperm characteristics following post thaw preparation showed no significant changes between the two groups (table 2). Conclusion: TYB and HSPM mediums are effective in cryoPreservation protocols when pre-freeze semen samples are used for Sperm preparation. Although there was a lack of statistically significant difference between the two groups, the post thaw motility patterns showed better values in samples frozen in TYB medium. Financial Support: None. Print this Page for Your Records Close WindowTable 1: Changes in pre-freeze and post-thaw semen characteristics after freezing with TYB and HSPMOasis, The Online Abstract Submission System Page 1 of 3http://www.abstractsonline.com/submit/SubmitPrinterFriendlyVersion.asp?ControlKey=%7... 5/1/2006
