Rheotaxis-based sperm separation using a biomimicry microfluidic device

Sperm selection is crucial to assisted reproduction, influencing the success rate of the treatment cycle and offspring health. However, in the current clinical sperm selection practices, bypassing almost all the natural selection barriers is a major concern. Here, we present a biomimicry microfluidic method, inspired by the anatomy of the female reproductive tract, that separates motile sperm based on their rheotaxis behavior to swim against the flow into low shear rate regions. The device includes micropocket geometries that recall the oval-shaped microstructures of the female fallopian tube to create shear protected zones for sperm separation. Clinical tests with human samples indicate that the device is capable of isolating viable and highly motile sperm based on their rheotaxis responses, resulting in a separation efficiency of 100%. The device presents an automated alternative for the current sperm selection practices in assisted reproduction.

EFFECT OF SWIM-UP AND GLASS WOOL TECHNIQUES, WITH ADDING ANTIOXIDANTS TO TRIS EXTENDER ON IMPROVING POST-CRYOPRESERVED TOTAL SPERM CHARACTERISTICS IN STRAW AND FREEZABILITY PERCENTAGE FOR LOW SEMEN QUALITY OF HOLSTEIN BULLS

This study was conducted to investigate the effect of swim-up and glass wool as sperm separation techniques with adding vitamin E and superoxide dismutase (SOD) to Tris extender on improving post-cryopreserved total sperm characteristics in straw and freezability percentage for low semen quality of Holstein bulls. Low and good of semen were extended using Tris extender. Good semen quality (GSQ) was divided into 3 groups (L1; Tris extender, L2; 2 mM vitamin E, L3; 200 IU SOD) Low semen quality (LSQ) was divided into two main groups, and subdivided into 3 sub-groups (L4; Tris extender, L5; 2 mM vitamin E, L6; 200 IU SOD). In the second main group, swim-up and glass wool techniques were used with adding vitamin E and SOD and subdivided into 3 sub-groups with each technique, and referred to L7, L8 and L9 for swim-up technique and L10, L11 and L12 for glass wool technique. Sperms with good motile intact acrosome and plasma membrane normal morphology of sperm were obtained using the glass wool separation technique with adding vitamin E.

Effect of Microfluidic Sperm Separation vs. Standard Sperm Washing Processes on Laboratory Outcomes and Clinical Pregnancy Rates in an Unselected Patient Population

A prospective, multicenter, randomized, sibling oocyte study was conducted with 86 couples to evaluate if a microfluidic sperm separation device improved ICSI sperm selection and subsequent cycle outcomes of fertilization, blastocyst utilization, ploidy, and clinical pregnancy rate when applied to a general patient population. Patients with at least 10 metaphase II oocytes were enrolled in the study and sibling oocyte groups were split in half. One half of the oocytes underwent ICSI with the control processed sperm and the other half were injected with sperm sorted by the ZyMōt microfluidic sperm separation device. Fertilization rate was recorded and resulting blastocysts were biopsied and evaluated for ploidy status with NGS. Euploid, non-mosaic embryos were randomly selected for single embryo transfer. A total of 787 oocytes were evaluated in the ZyMōt group and 777 in the control group. No statistical differences were observed between ZyMōt and control processing methods in any of the study outcomes evaluated. It is possible that the selection of normal, progressive sperm for ICSI, and the repair capacity of oocytes are sufficient to promote normal embryonic development in the general infertility population.

P–012 A new sperm preparation technique for ICSI

Study question Is there any difference in this technique(swim-in) according to clinical results in ICSI cycles? Summary answer Fertilization rates are significantly better with this new technique(swim-in). There is no difference in pregnancy and ongoing pregnancy rates between swim-in and DGS techniques. What is known already Sperm separation is crucial in assisted reproductive technologies, based on different principles like migration, filtration, or density gradient centrifugation techniques. However, there are some studies that centrifugation steps and using gradient solutions may increase sperm DNA fragmentation. On the other hand, multiple contacts to different plastic surfaces such as pipettes, plastic tubes, etc., and also pipetting the ejaculate in sperm preparation may have detrimental effects. The ideal sperm separation technique should be non-invasive, not time-consuming, easy, and cost-effective, not cause sperm damage or non-physiological alterations of the separated sperm cells, eliminate dead spermatozoa and other cells, including leukocytes. Study design, size, duration This is a prospective randomized study between 01/02/2019...01/01/2021. Sperms were prepared either with this new technique that we call “swim-in”(n = 359) or with density gradient centrifugation(n = 404) before microinjection (ICSI). Fertilization rates, clinical pregnancy rates, and on-going pregnancy rates were compared between groups. Sperm motility is less than%20 patients are not included in the study. t-test and Chi-square test used. p < 0.05 significant. Participants/materials, setting, methods For this new centrifugation free technique(swim-in), a Braun injector, 21G green needle, and sperm washing medium are used. First, we pulled a 0.5 ml sperm wash medium in a Braun injector. After sperm liquefaction, we put Braun injector with the needle in the sperm cap perpendicularly and waited 30 minutes at 37-degree Celcius. Sperms swam through the needle to the injector used for ICSI. For DGC sperms were prepared according to the 2015 WHO manual. Main results and the role of chance: We try to make a centrifugation-free, gentle, cost-effective, and easy sperm preparation technique and called it a swim-in technique. Groups were comparable according to mean age, collected oocyte numbers, and oocyte maturation ratios. Fertilization rates are%73,2(1786/2438) vs%66,9(2167/3237) respectively for swim-in and DGC groups. Clinical pregnancy rates are%45,1 vs%42,3 respectively for swim-in and DGC groups. On-going pregnancy rates are%41,5 vs%39,6 for swim-in and DGC respectively. There is a significant difference between the two techniques according to fertilization rates but no difference according to clinical pregnancy and ongoing pregnancy rates. Limitations, reasons for caution Sperms can move by their own motility from ejaculate to the sperm washing medium for that reason with severe sperm problem patients are not suitable for this technique. Wider implications of the findings: This technique is centrifugation free so there are no detrimental g-forces. The needle used may mimic the physiological environment like Uterine Tube. Alternative sperm washing mediums like follicular fluid or chemical attractor progesterone rich medium can be used.DNA fragmentation rates and live birth rates should be established for future aspect. Trial registration number Not applicable

Antibody-Conjugated Magnetic Beads For Sperm Sexing By A Multi-Wall Carbon Nanotube Microfluidic Device

Bull production industries demand specific sexual bulls in response to different business productivity requirements. Therefore, sexual separation of sperm is essential for the livestock industry, increasing the demand for assisted reproductive technologies (ARTs). A popular sexual separation method in ART is monoclonal antibody enhancement, which does not affect the quality of the sperm and has specificity with high sexual separation accuracy. Moreover, microfluidic technique is an innovative tool for sperm separation, with increased installation efficiency and convenience but low accuracy and speed of detection. In this study, particles were separated using magnetic beads, magnetic beads on monoclonal antibodies, and sperm with the function of magnetic beads on monoclonal antibodies by controlling the microfluidic system fluid flow rate at 2 µL/min and releasing a 2.5-V electric field. This microfluidic device separated 100% and 95.42% of the magnetic beads and magnetic beads on monoclonal antibodies, respectively. The sperm separation test results showed that positively charged sperm Y moved to the negative electrical fluid channel with a specially designed multi-walled carbon nanotube microfluidic device that showed an X- and Y-sperm separation with a success rate of 80.12%. Compared with natural pregnancy (X- and Y-sperm ration 50:50, 50%), our proposed method is approximately 30.12% better.

EFFECT OF SWIM-UP AND GLASS WOOL TECHNIQUES, WITH ADDING ANTIOXIDANTS TO TRIS EXTENDER ON IMPROVING POST-CRYOPRESERVED SOME SEMEN ATTRIBUTES OF LOW SEMEN QUALITY FOR HOLSTEIN BULLS

This study was conducted to investigate the effect of swim-up and glass wool as sperm separation techniques with adding vitamin E and superoxide dismutase (SOD) to Tris extender on improving some post-cryopreserved semen quality for Holstein bulls. Low and good of semen were extended using Tris extender. Good semen quality (GSQ) was divided into 3 groups (L1; Tris extender, L2; 2 mM vitamin E, L3; 200 IU SOD) Low semen quality (LSQ) was divided into two main groups, and subdivided into 3 sub-groups (L4; Tris extender, L5; 2 mM vitamin E, L6; 200 IU SOD). In the second main group, swim-up and glass wool techniques were used with adding vitamin E and SOD and subdivided into 3 sub-groups with each technique, and referred to L7, L8 and L9 for swim-up technique and L10, L11 and L12 for glass wool technique. Improving sperms’ cell motility and live sperm and reducing total sperms’ abnormalities percentages of low semen quality were noticed using swim-up and glass wool techniques were used with adding vitamin E and SOD. In conclusion, glass wool filtration separates out dead, abnormal and immotile sperm cells, a good sperm harvested by this technique.

Comparison of three sperm selection methods for ICSI-DGC, Cumulus column, and incubation with supernatant product of adipose tissue-derived adult stem cells: An experimental study

Background: The examination of sperm parameters and sperm DNA integrity are necessary for male fertility expression. These parameters can be affected by the method of sperm separation. Objective: To measure the damage of each sperm separation method on the sperm parameters and sperm DNA integrity. Materials and Methods: In this experimental study, semen samples of 20 infertile men with asthenoteratozoospermia (Infertility Research Center, Qom, Iran, 2017) were processed in three ways: density gradient centrifugation (DGC), cumulus column, and incubation with supernatant products of adipose tissue-derived adult stem cells (SPAS). The results of sperm parameters and DNA fragmentation before and after the process were statistically analyzed. Results: The number of separated sperms by normal morphologies during the SPAS and the cumulus column was significantly more than the corresponding population in the DGC group. In addition, although all three methods have the same ability to increase total sperm motility and the number of recovered sperms, in the field of forwarding movement and DNA fragmentation, the SPAS method performed more efficiently (p = 0.021). Conclusion: Sperm parameters and DNA fragmentation in the SPAS group were better than those in the DGC and cumulus column groups. Furthermore, it has been shown that the sperm capacity was increased with the SPAS method. However, the rearrangement of sperm chromatin by reducing the disulfide bridges and providing the possibility of re-histone over capacity causes a significant reduction in DNA fragmentation. Key words: Sperm, DNA fragmentation, Cumulus oophorus column, SPAS.

High efficiency rare sperm separation from biopsy samples in an inertial focusing device

Immotile and rare sperm isolation from a complex cell background is an essential process for infertility treatment. The traditional sperm collection process from a biopsy sample requires long, tedious searches,...