Introduction
sex-linked diseases
The desire to have offspring of a specific sex has a long history but has been particularly present since the 1970s with the early appearance of assisted reproduction. The reasons for choosing a child’s sex may be social, such as a desire for family balancing [Swim-up–based techniques allow spermatozoa selection without the use of centrifugation. For this method, the sperm specimen is placed in a tube, and a culture medium is carefully layered on top. In some instances, the tube is placed at a 45-degree angle at room temperature, or at 37°C. Spermatozoa are then allowed to swim up into the culture medium for approximately one hour. For female sex selection, the top layer is collected for use. For male sex selection, a modified technique is used where a small fraction of the top layer is first discarded, and the bottom portion is retrieved instead. However, there have been conflicting reports on the efficacy of this method. Some studies have observed a >70% sex enrichment [Unlike the swim-up selection method, which is dependent on sperm density, free flow electrophoresis selects for sex-specific spermatozoa according to net charge. A buffer is circulated through the chambers by electrodes while the semen specimen is deposited into the chamber continuously. This method was originally observed to provide a 90% enrichment for female and an 80% enrichment for male [Another approach, utilizing a Sephadex gel filtration, separates cells by size. Supposedly, lighter cells, such as male spermatozoa, are easily trapped, whereas those of larger molecular weight, X-bearing spermatozoa, can easily pass through. Therefore, although this technique has been shown to achieve an 80% accuracy for the proportion of female spermatozoa [The Percoll process employs discontinuous gradient layers at varying concentrations, where the semen specimen is layered on top of the gradients and centrifuged. This technique is effective at increasing overall sperm motility, but when applied for sperm sex selection, it has yielded inconsistent results [Ericsson’s Albumin Method is performed by layering aliquots of the sperm specimen on human serum albumin in a column, and then removing the individual layers at specific time intervals. An enrichment of female and male spermatozoa with 75% and 85% accuracy, respectively, was achieved with the use of this technique [Perhaps the most effective, yet controversial, method of sperm sex selection is flow cytometry. With this technique, sperm cells are exposed to fluorescent dyes that label their genetic material. Since X-bearing spermatozoa have 2.8% more genetic material, they fluoresce brighter, thereby differentiating them from Y-bearing spermatozoa. This method, capable of achieving a 90% enrichment for X- and 80% enrichment for Y-bearing spermatozoa [These methods all have advantages and limitations. There have been many variations of each technique, specifically related to the timing of centrifugation and the concentrations of gradient layers. Some methods, in addition to being time consuming, require costly equipment, such as a cell sorter. Furthermore, although there have been several reports on the efficacy of these techniques, there have also been studies disputing their effectiveness [Based on the limitations of these techniques, we propose a novel sperm sex selection method that appears to be consistently safe and effective. Moreover, since very few studies have assessed, for a single method, the technical efficacy of skewing a sperm specimen toward a specific sex, the clinical reliability of generating embryos of that desired sex, and the method’s impact on offspring health, we aimed to conduct a thorough evaluation of our novel method that addresses each of the aforementioned aspects in order to propose a valuable, feasible, and effective method to provide couples with a higher proportion of embryos of their desired sex. In addition to the medical rationale of avoiding sex-linked diseases or family balancing, the scientific rationale for this sperm-based sex selection technique is to introduce an ethical method to successfully skew the proportion of embryos towards a couple’s desired sex without impairing embryo developmental competence or offspring health, while minimizing embryo wastage. For couples undergoing intracytoplasmic sperm injection (ICSI), we performed a fluorescent in situ hybridization (FISH) assessment on spermatozoa pre- and post-selection to validate the efficacy of our technique. Our analytical plan also included an assessment of the proportion of the conceptuses’ sex to confirm clinical reliability. We also followed up on ICSI clinical outcomes and child/newborn health to establish the safety of this sex selection method.