Abstract
BackgroundThe morphology of spermatozoa is a fundamental aspect to consider in fertilization, sperm pathology, assisted reproduction and contraception. Head, neck, midpiece, principal and terminal part of flagellum are the main sperm components to investigate for identifying morphological features and related anomalies. Recently, scanning near-field optical microscopy (SNOM), which belongs to the wide family of nanoscopic techniques, has opened up new routes for the investigation of biological systems. SNOM is the only technique able to provide simultaneously highly resolved topography and optical images with a resolution beyond the diffraction limit, typical of conventional optical microscopy. This offers the advantage to obtain complementary information about cell surface and cytoplasmatic structures.ResultsIn this work human spermatozoa both healthy and with morphological anomalies are analyzed by SNOM, to demonstrate the potentiality of such approach in the visualization of sperm morphological details. The combination of SNOM topography with optical (reflection and transmission) images enables to examine typical topographic features of spermatozoa together with underlying cytoplasmic structures. Indeed the head shape and inner components as acrosome and nucleus, and the organization of mitochondria in the midpiece region are observed. Analogously for principal tract of the tail, the ridges and the columns are detected in the SNOM topography, while their internal arrangement can be observed in the corresponding SNOM optical transmission images, without requiring specific staining procedures or invasive protocols.ConclusionsSuch findings demonstrate that SNOM represents a versatile and powerful tool to describe topographical and inner structural details of spermatozoa simultaneously. This analysis could be helpful for better characterizing several morphological anomalies, often related to sperm infertility, which cannot be examined by conventional techniques all together.Electronic supplementary materialThe online version of this article (doi:10.1186/s12951-014-0061-5) contains supplementary material, which is available to authorized users.
Highlights
The morphology of spermatozoa is a fundamental aspect to consider in fertilization, sperm pathology, assisted reproduction and contraception
The potentialities of scanning near-field optical microscopy (SNOM) have been exploited to investigate the morphology of fixed human spermatozoa deriving from samples of patients with normozoospermia and teratozoospermia
In summary, we demonstrate that SNOM technique enables to describe the cell surface and the arrangement of the spermatozoa internal structures both in healthy and anomalous sperm by acquiring topographic and optical images simultaneously
Summary
The morphology of spermatozoa is a fundamental aspect to consider in fertilization, sperm pathology, assisted reproduction and contraception. SNOM is the only technique able to provide simultaneously highly resolved topography and optical images with a resolution beyond the diffraction limit, typical of conventional optical microscopy This offers the advantage to obtain complementary information about cell surface and cytoplasmatic structures. Conventional semen parameters such as sperm concentration, motility and morphology are generally used to assess male fertility [1,2,3]. Subdiffraction resolution optical images (reflection, transmission, back-reflection and fluorescence) and high-resolution (nanometer level) topographic image are simultaneously generated Such technique has been successfully applied for the investigation of cell membranes at molecular level [35,36,37,38], subcellular structures [39,40], thin-sections [41] and chromosomes [42]. We exploit the advantage of SNOM technique to describe simultaneously different superficial and internal morphological details of human spermatozoa from normozoospermia and teratozoospermia, without needing invasive or expensive sample preparation
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