Reconstruction of bovine spermatozoa substances distribution and morphological differences between Holstein and Korean native cattle using three-dimensional refractive index tomography

Abstract

Measurements of the three-dimensional (3D) structure of spermatozoon are crucial for the study of developmental biology and for the evaluation of in vitro fertilization. Here, we present 3D label-free imaging of individual spermatozoon and perform quantitative analysis of bovine, porcine, and mouse spermatozoa morphologies using refractive index tomography. Various morphological and biophysical properties were determined, including the internal structure, volume, surface area, concentration, and dry matter mass of individual spermatozoon. Furthermore, Holstein cows and Korean native cattle spermatozoa were systematically analyzed and revealed significant differences in spermatozoa head length, head width, midpiece length, and tail length between the two breeds. This label-free imaging approach provides a new technique for understanding the physiology of spermatozoa.

High-resolution 4-D acquisition of freely swimming human sperm cells without staining

“…This is a unique case among human cells. Other types of cells with rapid dynamics, such as cardiomyocytes, cannot be straightforwardly imaged by the proposed method but rather need to rely on rapid illumination rotation with limited scanning angle range [e.g., (18)]. Thus, the suggested method assumes progressive sperm cells with typical repetitive rotation of the sperm head, including continuous unidirectional rolling across 360°, a condition that is fulfilled for most of the cells, as we have observed from the acquired data.…”

“…To position the processed projections in the 3-D Fourier space, it is essential to know the angle at which the projection of the object was captured. Existing tomographic approaches are based on either illumination rotation (18) or controlled sample rotation (19), allowing a priori knowledge on the angles of the acquired interferometric projections. Alternatively, random rotation of cells while flowing along a microfluidic channel or perfusion chamber can be used for tomography for a degenerate case, assuming rotation around a single axis (20,21).…”

“…As the phase values of sperm cells often approach ~5 radians in a watery medium when tilted on their side, this condition is not met for all phase images. The Rytov approximation for the diffraction algorithm, on the other hand, is valid as long as the phase gradient in the sample is small (22,(37)(38)(39), as is generally the case for biological cells, and is thus a reasonable choice for applying tomography on sperm cells (18).…”

“…In tomographic phase microscopy, interferometric projections of the specimen taken from multiple angles are processed and positioned in the 3-D Fourier space so that the RI map can be reconstructed. Recently, a commercial tomographic microscope was used to recover the 3-D RI distributions of animal (nonhuman) sperm cells (18). Yet, this method scans only 30 illumination angles and at a limited angular range; thus, it is unable to fully image freely swimming sperm cells.…”

Editorial: Quantitative Phase Imaging and Its Applications to Biophysics, Biology, and Medicine

“…Recent developments in quantitative phase imaging (QPI) [4] have highly expanded the applicability of the refractive index as a reporter for advanced biological studies [5]. By directly and quantitatively measuring refractive index distributions or optical phase delay information, QPI provides various pieces of morphological and biophysical information about live cells and tissues, generating a series of new methods for cell biology [6,7], biophysics [8], reproductive science [9][10][11], infectious diseases [12], hematology [10,13], and neuroscience [14,15].…”