Ultrastructural Investigation of Human Sperm Using Atomic Force Microscopy

Abstract

Ultrastructural investigation of human sperm in its natural environment (without fixation, dehydration, embedding, sectioning, etc.) was carried out by using atomic force microscope (AFM) in its tapping mode. This technique permits the examination of fine structural details of undamaged sperm and its topography with precision. Moreover, it allows 3D reconstruction of images and enhances the contrast to resolve details such as mitochondria that surround the axoneme at the sperm middle piece. An organized structure has been found in the flageller axoneme region. Ultrastructure also reveals folding and details of the depression of the membrane that cannot be examined with conventional techniques.

The application of scanning near field optical imaging to the study of human sperm morphology

“…The rest of the flagellum in three points is measured as shown in Figure 3 A: profile 4, height of 120 nm; profile 5, height of 90 nm; profile 6, height of 70 nm. These results, in agreement with literature reported values, point out that the scanning of the spermatozoa by SNOM probe does not introduce artifacts and does not damage cell integrity [ 26 , 29 ].…”

“…Figure 5 A and B display the SNOM 3D representation (A) and topography (B) of the head. In Figure 5 C the head longitudinal profile provides a length of 5.2 μm, in agreement with the range of values obtained in literature with the size of integral sperm head [ 15 , 26 , 45 - 47 ]. Head appears flattened on the frontal part (250 nm on average in thickness) respect to the back (950 nm).…”

“…The profile points into evidence four bumps (indicated by the black arrows), which likely are due to the presence of mitochondria helicoidally surrounding axonema (Figure 2 ), as also observed in previous AFM work [ 26 ]. The total length of this region is found to be 4.5 μm in accordance with the range of values obtained in literature [ 26 ]. In Figure 3 E horizontal cross section profiles taken on the different part of the sperm are shown (white arrows in Figure 3 A).…”

“…The analysis of the topographical image is shown in Figure 3 D and E. Figure 3 D shows a longitudinal profile of the midpiece, between the head and anulus along the red arrow direction of Figure 3 A. The profile points into evidence four bumps (indicated by the black arrows), which likely are due to the presence of mitochondria helicoidally surrounding axonema (Figure 2 ), as also observed in previous AFM work [ 26 ]. The total length of this region is found to be 4.5 μm in accordance with the range of values obtained in literature [ 26 ].…”

“…Recently scanning probe microscopy (SPM) techniques have been exploited to improve the knowledge of spermatozoa morphology. Particularly, atomic force microscopy (AFM) has allowed investigating the topography of spermatozoa with nanometric resolution and without needing invasive sample preparation [ 26 - 29 ]. Moreover, either fixed or living cells can be imaged by this technique.…”