Abstract
Infertility is a widespread problem, and in some cases, the routine basic semen analysis is not sufficient to detect the cause of male infertility. The use of the scanning electron microscope (SEM) could provide a detailed insight into spermatozoa morphology, but it requires specific sample preparation techniques. The purpose of this study was to select, adjust, and optimize a method for the preparation of spermatozoa samples prior to SEM analysis, and to establish the protocol required for its use in clinical practice. We examined sperm samples of 50 men. The samples were fixed with modified iso-osmolar aldehyde solution followed by osmium post-fixation. In the first method, dehydration of the cells and subsequent critical point drying (CPD) were performed on a coverslip. In the second method, the samples were dehydrated in centrifuge tubes; hexamethyldisilazane (HMDS) was used as a drying agent instead of CPD, and the samples were air-dried. The third procedure was based on a membrane filter. The samples were dehydrated and dried with HMDS in a Gooch crucible, continuously, without centrifugation or redispersion of the sample. Our results showed that the fixation with modified iso-osmolar aldehyde solution followed by osmium post-fixation, and combined with dehydration and CPD on a coverslip, is the most convenient procedure for SEM sample preparation. In the case of small-size samples or low sperm concentration, dehydration and drying with HMDS on the membrane filter enabled the best reliability, repeatability, and comparability of the results. The presented procedures are suitable for routine use, and they can be applied to confirm as well as to correct a diagnosis.
Highlights
Poor quality of semen is a common cause of male infertility [1]
scanning electron microscope (SEM) analysis of spermatozoa prepared on coverslips and critical point drying (CPD)
CPD enables the fluid to transform to the gaseous phase without any phase boundary, meaning that there are no forces of surface tension that could damage the structure of the sample (Figure 1A and B)
Summary
Poor quality of semen is a common cause of male infertility [1]. In one out of five couples who are having fertility problems, there is an issue with the male partner [2,3]. The semen analysis or spermiogram includes physical, morphological, and biochemical analysis of the ejaculate. All semen parameters should be considered [4]. The semen analysis is important in determining the fertilizing potential of spermatozoa [5,6,7]. Spermiogram is a part of andrological examination and currently remains the most appropriate test for assessing the male fertility [8]
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
