Sperm Sorting Research Articles

Towards a better testicular sperm extraction: novel sperm sorting technologies for non-motile sperm extracted by microdissection TESE.

Non-obstructive azoospermia (NOA) is the most severe form of male factor infertility. It is characterized by a lack of spermatogenesis in the seminiferous tubules. Microdissection testicular sperm extraction (microTESE) has significantly improved testicular sperm retrieval rates compared to conventional techniques for NOA. Following testicular biopsy, the sperm is usually non-motile and contained within seminiferous tubules requiring extensive laboratory processing to find individual sperm sufficient for artificial reproductive technologies (ART). Current techniques include mechanical and enzymatic processing which is time-consuming and often damaging to sperm. We review novel techniques that may help improve sperm retrieval rates after microTESE including microfluidics (dielectrophoretic cell sorting, spiral channel sorting, and pinched flow fractionation), fluorescence-activated cell sorting (FACS), and magnetic-activated cell sorting (MACS).

Sperm Elde Etmek için Hangisini Seçmeliyiz? Kardeş Oosit Çalışması: Yüzdürme Yöntemine Karşı Yoğunluk Gradyan

Sperm Elde Etmek için Hangisini Seçmeliyiz? Kardeş Oosit Çalışması: Yüzdürme Yöntemine Karşı Yoğunluk Gradyan

The use of sexed semen in dairy herds

Artificial insemination of cattle became a commercial reality in 1936, and in 1952 the first calves were born as a result of freeze-thawed semen insemination. The long-held aspiration for pre determination of sex became a reality with the development of sperm sorting technology with the first female calves being born in the UK in 1999. While most breeding companies supply sexed semen sorted by the original flow cytometry method, Genus ABS have developed the method of sperm deactivation. Whatever the method sexing accuracy exceeds 90% and conception rates can be expected to be comparable to conventional semen but only when management is good and importantly there are considerable variations in the fertility of different bulls as a result of the sorting process. The use of sexed semen offers producers an opportunity to maximise the beef production from the dairy herd while accelerating genetic improvement.

Comparison of microfluid sperm sorting chip and density gradient methods for use in intrauterine insemination cycles

To compare the effect of microfluiding sperm sorting chip and density gradient methods on ongoing pregnancy rates (PRs) of patients undergoing IUI. Retrospective cohort study. Hospital IVF unit. Couples with infertility undergoing IUI cycles between 2017 and2018. Not applicable. Ongoing PRs. A total of 265 patients were included in the study. Microfluid sperm sorting and density gradient were used to prepare sperm in 133 and 132 patients, respectively. Baseline spermiogram parameters, including volume, concentration, motility, and morphology, were similar between the two groups. Total motile sperm count was lower in the microfluiding sperm sorting group at baseline (35.96 ± 37.69 vs. 70.66 ± 61.65). After sperm preparation sperm motility was higher in the microfluid group (96.34 ± 7.29 vs. 84.42 ± 10.87). Pregnancy rates were 18.04% in the microfluid group and 15.15% in the density gradient group, and ongoing PRs were 15.03% and 9.09%, respectively. After using multivariable logistic regression and controling for confounding factors, there was a significant increase in ongoing PRs in the microfluid sperm sorting group. The adjusted odds ratio for ongoing pregnancy in the microfluid group compared with the density gradient group was 3.49 (95% confidence interval 1.12-10.89). The microfluid sperm sorting method significantly increased the ongoing PRs compared with the density gradient group in IUI cycles.

Does the use of microfluidic sperm sorting for the sperm selection improve IVF success rates in male factor infertility?

Does the use of microfluidic sperm sorting for the sperm selection improve IVF success rates in male factor infertility?

Does a microfluidic chip for sperm sorting have a positive add-on effect on laboratory and clinical outcomes of intracytoplasmic sperm injection cycles? A sibling oocyte study.

The most recent technologies for sperm sorting involve microfluidics. However, the most important question whether their use is of any advantage in terms of laboratory and clinical IVF/ICSI outcomes still remains controversy. In this study, we aimed to investigate whether a microfluidic sperm sorting device (Fertile Plus® ) has a positive add-on effect on laboratory and clinical outcomes. Sibling oocytes of 81 patients were assigned to two sperm sorting groups including swim up and Fertile Plus® . All embryos were cultured until day 5/6. Fertilisation, embryo quality and blastocyst development were assessed as primary outcomes among 81 patients; clinical pregnancy, implantation and live birth rates were analysed as secondary outcomes as a subgroup analysis due to transfer cancellations. No statistically significant differences were found between groups in terms of all outcomes analysed in laboratory and clinical terms (p>.05 for all). The results of this study suggest that sorting spermatozoa through Fertile chip does not improve laboratory outcomes significantly and does not seem to have a positive contribution to clinical outcomes.

A treatment approach for couples with disrupted sperm DNA integrity and recurrent ART failure

ObjectiveTo test a novel method to select spermatozoa with high chromatin integrity.DesignSpecimens with high sperm chromatin fragmentation (SCF) were selected by density gradient selection (DGS) and microfluidic sperm sorting (MSS).SettingAcademic medical center.Patient(s)Ejaculates from consenting men were processed by DGS/MSS. Couples underwent ICSI cycles with spermatozoa processed by DGS/MSS. Clinical outcomes were evaluated after embryo transfer.Intervention(s)SCF was measured by TUNEL. ICSI with spermatozoa selected by DGS and MSS was performed.Main outcome measure(s)Fertilization, embryo implantation, and pregnancy outcomes were compared between DGS and MSS.Result(s)A total of 23 men had an average SCF of 20.7 ± 10%. After DGS and MSS, the SCF was 12.5 ± 5% and 1.8 ± 1%, respectively. In couples who underwent ICSI, the average SCF was 28.8 ± 9%, which fell to 21.0 ± 9% after DGS and 1.3 ± 0.7% after MSS. Four couples underwent 11 ICSI cycles with DGS and achieved one (25%) pregnancy that resulted in pregnancy loss. In four subsequent ICSI cycles with MSS, an ongoing clinical pregnancy rate of 50% was achieved. Five additional couples underwent 12 cycles of ICSI with DGS. After preimplantation genetic testing for aneuploidy, 30.3% of the embryos were euploid. One pregnancy was achieved, resulting in pregnancy loss. With MSS, 31.5% of the embryos were euploid and 4 couples obtained a pregnancy. Finally, sixteen couples underwent 20 ICSI cycles solely with MSS at our center. Of these couples, 8 had failed 13 ICSI cycles with DGS elsewhere. These couples achieved an overall implantation of 34.5% (10/29) and a pregnancy rate of 58.8% (10/17).Conclusion(s)Microfluidic selection yielded spermatozoa with optimal genomic integrity and improved chances of obtaining a euploid conceptus.

Review on Sperm Sorting Technologies and Sperm Properties toward New Separation Methods via the Interface of Biochemistry and Material Science.

Successful fertilization in mammals requires spermatozoa to efficiently traverse the female reproductive tract to meet the egg. This process naturally selects high quality sperm cells for fertilization, but when artificial reproductive technologies are used such as in vitro fertilization, intracytoplasmic sperm injection, or intrauterine insemination, other methods of sperm selection are required. Currently, technology enables sperm sorting based on motility, maturity as defined by zeta potential or hyaluronic acid binding site expression, absence of apoptotic factors, appropriate morphology, and even sex. This review summarizes current knowledge on all known methods of sperm cell sorting, compares their efficiency, and discusses the advantages and limitations of each technique. Scope for further refinement and improvement of current methods are discussed as is the potential to utilize a variety of materials to innovate new methods of sperm separation.

Gravity and rheotaxis based sperm sorting device employing a cam-actuated pipette mechanism.

Until now, a swim-up or microchip-based method has been mainly utilized for separating normal sperm for use in assisted reproductive technology. However, it requires excessive sorting time due to preprocessing and collects a limited number of motile sperms. To improve this process, we propose a gravity-fed high motility sperm sorting device that utilizes the rheotaxis of sperm, which minimizes separation time and improves throughput. The device features a mesoscale microfluidic channel to maximize the throughput, and an outlet at the bottom is configured to control the fluid velocity in the channel by using gravity. To control and automate semen injection and suction of the sorted sperm, a pipette controller using a cam was fabricated. After constructing the system, a sorting experiment was performed using canine semen to confirm the separation efficiency. After injecting the semen in the channel, the delay time between injection and suction was measured and the relative improvement of the index of motility was investigated according to measured delay time. As a result of repeated experiments, it was confirmed that the highest improvement was obtained at a delay time of 80 s, and the mean velocity, %motility, MI, and motile sperm rates were improved by 8.94%, 32.58%, 35.48%, and 21.99%, respectively.

To Study the Effect of Magnetic Activated Sperm Sorting on the Outcome of in Vitro Fertilization-embryo Transplantation

To Study the Effect of Magnetic Activated Sperm Sorting on the Outcome of in Vitro Fertilization-embryo Transplantation

The Evaluation of the Effect of Microfluidic Sperm Sorting Chip 'Labs-on-a-chip' on IVF Success in Male Factor

The Evaluation of the Effect of Microfluidic Sperm Sorting Chip 'Labs-on-a-chip' on IVF Success in Male Factor

Rheotaxis Based High-Throughput Motile Sperm Sorting Device

A progressive motile sperm sorting device with a diffuser-type microfluidic channel is proposed to improve fertilization in assisted reproductive technologies. When sperm cells swim against a fluid flow, they are positioned according to their motility in specific channel zones. A numerical simulation is carried out in a meso-sized diffuser-type channel which can realize high throughput to verify the working principle and the sorting device is fabricated based on the simulation result. Furthermore, to analyze the condition of the semen and investigate the experimental results, a novel image processing method is proposed. With the proposed analyzing technique, the sperm sorting experiment is performed in the channel using the canine semen sample. As a result, the %motility increases from 59.89 to 69.36%, however, it does not satisfy the reported guideline for canine sperm motility (%motility > 70%). Therefore, we perform a washing process by injecting DPBS fluids to improve indexes related to motility. After double washing procedures, we could increase %motility from 59.89 to 82.24%, motile sperm rate from 33.38 to 53.10% and motility index from 26.35 to 49.44 in Zone A, comparing to the prepared sample.

Microfluidic technology for in vitro fertilization (IVF)

With the recent development in science and technology, the advancement in in vitro fertilization to treat infertility has been one of the most revolutionary advances. However, the success rate of the IVF process depends on the efficiency of each step of the process. The physical tools used to enhance the process continue to change. Microfluidics technology is an emerging technology being used in multiple biological applications for the miniaturization and specification of laboratory techniques. Technology is used along with IVF to enhance the outcome by facilitating every step of the process. Microfluidics can be used to handle gametes, culture embryos, cryopreservation, and for many other applications. This review will highlight the applications of microfluidics in different stages of IVF, including the handling of gametes, sperm collection and isolation, sperm sorting, embryo culture, cryopreservation and the fabrication process of microfluidics, focusing on the benefits and shortcomings of these applications.

Design Integrated Microfluidic Chips for Clinical Applications

A biosensor is generally defined as an analytical device, which converts the biochemical responses into the quantifiable electronic signals. Its extensive utilities in medical diagnosis, toxicity testing, chemical analysis, food industry, and many other areas for quantitative assessments has appeared. Quartz crystal microbalance and field-effect transistor based devices have been demonstrated to possess exceptional characteristics and outstanding performance while conjugated with the advanced nanotechnology. However, both devices still have to face the nonspecific binding issue, which is a headache for researchers on all biosensor platforms. In our research, we have efficaciously reduce 95% nonspecific binding via immobilize the zwitterionic layer on the surface of the chip. Moreover, we integrate microfluidics, nanoparticles, rapid surface modification on several bio-sensing platforms to efficiently pretreat specimens and detect target biomolecules. In addition, a series advanced microfluidic ships are designed for lymphocyte isolation, active sperm sorting, and nanoparticle synthesis. Figure 1

(Invited) Design Integrated Microfluidic Chips for in Vitro Diagnosis

A biosensor is generally defined as an analytical device, which converts the biochemical responses into the quantifiable electronic signals. Its extensive utilities in medical diagnosis, toxicity testing, chemical analysis, food industry, and many other areas for quantitative assessments has appeared. Quartz crystal microbalance and field-effect transistor based devices have been demonstrated to possess exceptional characteristics and outstanding performance while conjugated with the advanced nanotechnology. However, both devices still have to face the nonspecific binding issue, which is a headache for researchers on all biosensor platforms. In our research, we have efficaciously reduce 95% nonspecific binding via immobilize the zwitterionic layer on the surface of the chip. Moreover, we integrate microfluidics, nanoparticles, rapid surface modification on several bio-sensing platforms to efficiently pretreat specimens and detect target bio-molecules. In addition, a series advanced microfluidic ships are designed for lymphocyte isolation, active sperm sorting, and nanoparticle synthesis. Figure 1

Distinct and independent dielectrophoretic behavior of the head and tail of sperm and its potential for the safe sorting and isolation of rare spermatozoa.

Often, in semen samples with minute amounts of sperm, even the single spermatozoon required to fertilize an oocyte cannot be found in the ejaculate. This is primarily because currently, sperm is generally searched for manually under a microscope. In this study, dielectrophoresis (DEP) was investigated as an alternative automated technique for sorting sperm cells. Using a quadrupolar electrode array it was shown that the head and tail of the sperm had independent and unique crossover frequencies corresponding to the transition of the DEP force from repulsive (negative) to attractive (positive). These surprising results were further analyzed, showing that the head and tail have their own distinct electrical properties. This significant result allows for the sperm's head, which contains the DNA, to be distanced from potentially damaging high electric fields using negative DEP while simultaneously manipulating and sorting the sperm using the positive DEP response of the tail. A proof of concept sorting chip was designed and tested. The low crossover frequency of the tail also allows for the use of a higher conductivity, and thus more physiological, medium than the conventional DEP solutions. Although more research is required to design and optimize an efficient, user-friendly, and high-throughput device, this research is a proof of concept that DEP has the potential to automate and improve the processing of semen samples, especially those containing only rare spermatozoa.

What research is needed to improve commercial pig reproduction

Artificial insemination with fresh or stored semen is currently the only sperm technology used at a commercial scale in the pig industry. Attention should therefore be given for further improvement of the functionality and fertilizing ability of cryopreserved semen, as well as for sperm sorting for gender pre-selection. During the last two decades various proteins and polypeptides have been identified in boar seminal plasma, and the relevance of some of them to reproductive technologies has been discussed at this conference. The long- term goal should be to isolate/synthetize those seminal plasma proteins proven important for the spermatozoa, and use them as ingredients in media used for e.g. cryopreservation and sex sorting of semen. A close cooperation between biochemists, molecular biologists and animal scientists is necessary to reach this goal.

Pregnancy rates in cattle with cryopreserved sexed spermatozoa: effects of laser intensity, staining conditions and catalase

The overall aim of this research was to improve fertility of cattle inseminated with sexed spermatozoa by improving sperm sorting procedures. Six field trials were conducted in which 4,264 heifers were inseminated into the uterine body with cryopreserved sexed or unsexed control spermatozoa. Pregnancy or calving rates with doses of 2 x 10(6) sexed spermatozoa ranged from 32 to 51%; these averaged 69% of the pregnancy rates with 20 x 10(6) unsexed, control spermatozoa (range 53 to 79% of controls). Fertility of sexed spermatozoa was especially low on farms where control fertility was low. Accuracy of sexing ranged from 86 to 91%. Laser power of 150 mW for interrogating spermatozoa did not result in lower pregnancy rates (43%) than when power was decreased as much as possible for a particular sorting batch (50 to 130 mW) to still achieve sexing accuracy (38% pregnant). Addition of catalase to fluids containing spermatozoa was beneficial when thawed spermatozoa were incubated in vitro for 2 h but had no effect on pregnancy rates. There also was no effect on pregnancy rates between two concentrations of Hoechst 33342 for staining spermatozoa. Freezing 2 x 10(6) sexed spermatozoa at 20 x 10(6)/ml resulted in a slightly higher rate of pregnancy (P < 0.05) than at 10 x 10(6)/ml. The information obtained in these trials, along with other improvements, notably lowering pressure in the sorting system from 50 to 40 psi, has been used to improve procedures for sexing spermatozoa commercially.

MP52-06 MICROFLUDIC SPERM SORTING BY USING MICROCHIP AS A NEW METHOD FOR REPATED IVF FAILURE CASES: COMPARISON ITS’ EFFECT WITH CONTROL GROUP AND PRELIMINARY RESULTS

INTRODUCTION AND OBJECTIVES:Microfludic sperm sorting is a new method of sperm selection for ART procedures. The aim of the study was to evaluate the efficacy of this method for sperm sorting in in...

Use of microfluidics to sort stallion sperm for intracytoplasmic sperm injection

We determined if microfluidic sorting (MF) of frozen-thawed stallion sperm improves sperm population characteristics and results in embryo development after intracytoplasmic sperm injection (ICSI). The efficiency and efficacy of MF sperm separation was evaluated by comparing pre- and post-separation sperm population variables. Procedural comparisons were performed after sorting with MF, single-layer colloidal centrifugation (SLC) or swim-up (SU), and cleavage and embryo development were evaluated after ICSI using MF-sorted sperm. In Experiment 1, when compared to the original sperm sample, MF sorting resulted in a sperm subpopulation with greater motility, morphology, viability, and membrane as well as DNA integrity. After sorting by MF, SLC and SU in Experiment 2, motility, viability, and membrane integrity were similar for sperm sorted using MF and SLC; however, morphology and DNA integrity were greater in sperm sorted using MF when compared with SLC. Swim-up was the least effective sorting method. In Experiment 3, sperm were processed using MF and SLC prior to ICSI. Motility, morphology and DNA integrity were similar for sperm subpopulations sorted using either method; but viability was greater for sperm sorted using MF than SLC. Sorting did not improve sperm membrane integrity. Sorting with MF prior to ICSI resulted in similar cleavage and blastocyst development rates as SLC. We concluded that MF separation of stallion sperm resulted in a subpopulation with improved sperm population parameters, comparable or better than SLC and SU. Embryos were produced after ICSI using MF sperm sorting.