Sperm Preparation Techniques for Artificial Insemination - Comparison of Sperm Washing, Swim Up, and Density Gradient Centrifugation Methods

Natali, Ilaria

doi:10.5772/17026

Cited by 12 publications

(9 citation statements)

References 29 publications

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“…The efficiency of sperm preparation methods to separate bacteria from the seminal fluid and to avoid possible negative effects of bacteriospermia on the sperm recovery rates are essential aspects of a successful sperm handling procedure [30]. The choice to process ejaculates prior to their use in artificial insemination (AI) or in vitro fertilization (IVF) is currently supported by studies in roosters [31] rabbits [31,32] and boars [31,33], where the semen samples often suffer from a significant contamination which may severely affect the final quality of the sample and subsequently the fertilization success.…”

Section: Discussionmentioning

confidence: 99%

Antibiotics Versus Natural Biomolecules: The Case of In Vitro Induced Bacteriospermia by Enterococcus Faecalis in Rabbit Semen

et al. 2019

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Male subfertility is a global issue in human reproduction as well as in animal reproduction. Bacterial infection and semen contamination are still widely overlooked. As the collection of ejaculates is not a sterile process, it is necessary to add antimicrobial agents to avoid a possible depreciation of semen samples. As traditionally used antibiotics have been questioned because of an ever-increasing bacterial resistance, natural bioactive molecules could offer an alternative because of their antibacterial and antioxidant properties. As such, we decided to compare the effects of selected natural biomolecules (resveratrol-RES, quercetin-QUE and curcumin-CUR) with routinely used antibiotics in animal biotechnologies (penicillin-PEN, gentamicin-GEN and kanamycin-KAN) on the rabbit sperm vitality in the presence of Enterococcus faecalis. Changes in the sperm structural integrity and functional activity were monitored at 0, 2, 4 and 6 h. Computer-assisted sperm analysis (CASA) was used for the assessment of spermatozoa motility. Production of reactive oxygen species (ROS) was evaluated using chemiluminiscence, while the mitochondrial membrane potential (ΔΨm) was examined using the JC-1 dye. Finally, the sperm chromatin dispersion (SCD) test was used to assess DNA fragmentation, and changes to the membrane integrity were evaluated with the help of annexin V/propidium iodide. The motility assessment revealed a significant sperm motility preservation following treatment with GEN (p < 0.001), followed by PEN and CUR (p < 0.01). QUE was the most capable substance to scavenge excessive ROS (p < 0.001) and to maintain ΔΨm (p < 0.01). The SCD assay revealed that the presence of bacteria and antibiotics significantly (p < 0.05) increased the DNA fragmentation. On the other hand, all bioactive compounds readily preserved the DNA integrity (p < 0.05). In contrast to the antibiotics, the natural biomolecules significantly maintained the sperm membrane integrity (p < 0.05). The microbiological analysis showed that GEN (p < 0.001), KAN (p < 0.001), PEN (p < 0.01) and CUR (p < 0.01) exhibited the strongest antibacterial activity against E. faecalis. In conclusion, all selected biomolecules provided protection to rabbit spermatozoa against deleterious changes to their structure and function as a result of Enterococcus faecalis contamination. Therefore, administration of RES, QUE and/or CUR to rabbit semen extenders in combination with a carefully selected antibacterial substance may be desirable.

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Section: Discussionmentioning

confidence: 99%

Antibiotics Versus Natural Biomolecules: The Case of In Vitro Induced Bacteriospermia by Enterococcus Faecalis in Rabbit Semen

et al. 2019

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“…The sperms isolated with the density gradient centrifugation are not damaged by the ROS but with low DNA integrity. 7 Another study showed that the sperms which are obtained through twolayer density gradient centrifugation provide spermatozoa with a higher quality in terms of the motility, viability, and low DNA fragmented as compared to those which are obtained by the other conventional sperm preparation methods. 8 A comparative study of two-layer density gradient technique and swim-up method showed that the outcome of two-layer density technique is better than the swim-up method.…”

Section: Comparison Between Two-layer Density Gradient and Three-layementioning

confidence: 99%

Comparison between Two-Layer Density Gradient and Three-Layer Density Gradient Technique for Sperm Preparation at Aster Fertility Clinic, Dr. Hasan Sadikin General Hospital

Astarto

Tjahyadi

Jatnikasari

2014

ijihs

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Objective: To compare the efficacy of sperm preparation techniques modified by two layer density gradient technique (80%, 40%). Methods: Sperm preparation was performed by two-and three-layer density gradient technique by centrifugation at 500 rpm for 20 minutes using SpermGrad (Vitrolife, Sweden). Results: The average concentration of total sperm preparation by two-layer density gradient was 23.19 million with a standard deviation of 11.9 million while the result of three-layer density gradient preparation obtained a mean total sperm concentration of 19.64 million with a standard deviation of 10.033 million. The mean total concentration of sperm in three-layer was lower than two-layer density gradient with p <0.001. Conclusions: This study concludes that there was a significant difference between the concentration of sperm prepared using two-layer and three-layers density gradient technique.

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“…Namun, pada sampel dengan kualitas yang suboptimal seperti pada pasien infertil maka teknik density gradient centrifugation lebih sering digunakan. 11 Penelitian mengenai efektivitas preparasi semen sebelum proses kriopreservasi telah melaporkan hasil yang beragam. Terdapat dua protokol kriopreservasi berkaitan dengan preparasi semen, yakni protokol direct freezing dan protokol preparasi pra-freezing.…”

Section: Pendahuluanunclassified

“…Preparasi semen merupakan suatu teknik seleksi yang memiliki tujuan menuai populasi spermatozoa fungsional dari cairan ejakulat. 11,15,16 Meskipun terdapat beberapa teknik preparasi semen, teknik swim up dan DGC merupakan dua teknik yang umum digunakan. Pemilihan teknik preparasi semen disesuaikan dengan kualitas spermatozoa dalam cairan ejakulat.…”

Section: Pembahasanunclassified

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Density Gradient Centrifugation Pra-freezing Mengoptimalkan Persentase Morfologi Normal Spermatozoa Pasca-thawing

Suyono

Hinting²,

Lunardhi³

et al. 2018

mkb

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Kriopreservasi akan mengganggu struktur dan fungsi spermatozoa. Preparasi semen mampu menghasilkan spermatozoa dengan kualitas baik. Penelitian ini bertujuan menganalisis pengaruh preparasi semen dengan density gradient centrifugation (DGC) pra-freezing terhadap kualitas spermatozoa pasca-thawing. Penelitian dilakukan di boratorium Biologi Kedokteran, Fakultas Kedokteran Universitas Airlangga periode November 2017-Januari 2018. Penelitian eksperimental laboratoris dilakukan menggunakan cairan ejakulat volunter pria infertil. Semua sampel dibagi menjadi dua bagian, kelompok kontrol serta kelompok perlakuan berupa preparasi mini DGC. Setelah penambahan krioprotektan, selanjutnya dilakukan freezing. Pemeriksaan kualitas spermatozoa, meliputi motilitas, viabilitas serta persentase morfologi normal menggunakan metode WHO 2010, baik pra-freezing maupun pasca-thawing. Persentase perubahan kualitas spermatozoa pasca-thawing kedua kelompok dibandingkan menggunakan uji t dan bermakna jika nilai p<0,05. Total 20 sampel cairan ejakulat digunakan dalam penelitian ini. Persentase penurunan motilitas progresif, motilitas total, serta viabilitas pascathawing antara kedua kelompok tidak berbeda bermakna dengan nilai p masing-masing 0,422; 0,873 serta 0,432. Namun, penurunan persentase morfologi normal pasca-thawing pada kelompok kontrol jauh lebih besar daripada kelompok perlakuan dengan nilai p<0,001. Penelitian ini menyimpulkan bahwa preparasi semen berupa mini DGC pra-freezing mampu menghasilkan spermatozoa pasca-thawing dengan persentase morfologi normal yang lebih baik daripada protokol direct freezing.

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Sperm Preparation Techniques for Artificial Insemination - Comparison of Sperm Washing, Swim Up, and Density Gradient Centrifugation Methods

Cited by 12 publications

References 29 publications

Antibiotics Versus Natural Biomolecules: The Case of In Vitro Induced Bacteriospermia by Enterococcus Faecalis in Rabbit Semen

Antibiotics Versus Natural Biomolecules: The Case of In Vitro Induced Bacteriospermia by Enterococcus Faecalis in Rabbit Semen

Comparison between Two-Layer Density Gradient and Three-Layer Density Gradient Technique for Sperm Preparation at Aster Fertility Clinic, Dr. Hasan Sadikin General Hospital

Density Gradient Centrifugation Pra-freezing Mengoptimalkan Persentase Morfologi Normal Spermatozoa Pasca-thawing

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