Search

Cryopreservation of sperm is an essential technique in assisted reproduction in cattle. The objective of the study was to systematically review and synthesize the literature on bull semen quality evaluation based on the comparison of morphological and metabolic parameters of cryopreserved bovine spermatozoa such as DNA integrity, mitochondrial status, plasma membrane alterations, total motility, and morphology (% of abnormal cells). The electronic databases PubMed, Web of Sciences, Scopus, and Google Scholar were searched up to December 2023. Studies and references were included if they reported the following parameters: DNA integrity, mitochondrial status, plasma membrane alterations, total motility, and morphological aberrations (% of abnormal cells) for conventional cryopreserved bovine spermatozoa. After an electronic search, out of 1,526 original studies, only 40 were included in the meta-analysis. Standardized mean differences (SMD) with 95% confidence intervals were estimated for the chosen studies, and a meta-analysis was performed using a random effects model. The tau-squared (tau2) and inconsistency index (I2) quantified heterogeneity among different studies. The regression analysis for the evaluated parameters showed a positive correlation between mitochondrial membrane potential (MMP), total motility, and abnormal morphology and a negative correlation between DNA fragmentation index (DFI) and total motility and MMP. Moreover, subgroup analysis demonstrated similar associations for dairy and non-dairy bull breeds, albeit with lower I2 values. The presence of publication bias was confirmed by Egger's test, except for the MMP parameter. A multi-parametric analysis of morphological and metabolic parameters can address the existing limitations of cryopreserved bovine spermatozoa quality assessment. Combining imaging flow cytometry (IFC) with standardization of sperm pre-processing and optimization of the experimental protocols may help to differentiate sperm from cellular debris and cytoplasmic droplets of similar size and alleviate limitations demonstrated by conventional sperm analysis. [ABSTRACT FROM AUTHOR]

Multi-nuclearity is a common feature for cells in different cancers. Also, analysis of multi-nuclearity in cultured cells is widely used for evaluating the toxicity of different drugs. Multi-nuclear cells in cancer and under drug treatments form from aberrations in cell division and/or cytokinesis. These cells are a hallmark of cancer progression, and the abundance of multi-nucleated cells often correlates with poor prognosis.The use of standard bright field or fluorescent microscopy to analyze multi-nuclearity at the quantitative level is laborious and can suffer from user bias. Automated slide-scanning microscopy can eliminate scorer bias and improve data collection. However, this method has limitations, such as insufficient visibility of multiple nuclei in the cells attached to the substrate at low magnification.Since quantification of multi-nuclear cells using microscopic methods might be difficult, imaging flow cytometry (IFC) is a method of choice for this. We describe the experimental protocol for the preparation of the samples of multi-nucleated cells from the attached cultures and the algorithm for the analysis of these cells by IFC. Images of multi-nucleated cells obtained after mitotic arrest induced by taxol, as well as cells obtained after cytokinesis blockade by cytochalasin D treatment, can be acquired at a maximal resolution of IFC. We suggest two algorithms for the discrimination of single-nucleus and multi-nucleated cells. The advantages and disadvantages of IFC analysis of multi-nuclear cells in comparison with microscopy are discussed., (© 2023. The Author(s).)

Microcystis is a globally known cyanobacterium causing potentially toxic blooms worldwide. Different morphospecies with specific morphological and physiological characters usually co-occur during blooming, and their quantification employing light microscopy can be time-consuming and problematic. A benchtop imaging flow cytometer (IFC) FlowCam (Yokogawa Fluid Imaging Technologies, USA) was used to identify and quantitate different Microcystis morphospecies from environmental samples. We describe here the FlowCam methodology for sample processing and analysis of five European morphospecies of Microcystis common to the temperate zone. The FlowCam technique allows detection of different Microcystis morphospecies providing objective qualitative and quantitative data for statistical analysis., (© 2023. Springer Science+Business Media, LLC, part of Springer Nature.)

Fluorescence methods are widely used for the study of marine and freshwater phytoplankton communities. However, the identification of different microalgae populations by the analysis of autofluorescence signals remains a challenge. Addressing the issue, we developed a novel approach using the flexibility of spectral flow cytometry analysis (SFC) and generating a matrix of virtual filters (VF) which allowed thorough examination of autofluorescence spectra. Using this matrix, different spectral emission regions of algae species were analyzed, and five major algal taxa were discriminated. These results were further applied for tracing particular microalgae taxa in the complex mixtures of laboratory and environmental algal populations. An integrated analysis of single algal events combined with unique spectral emission fingerprints and light scattering parameters of microalgae can be used to differentiate major microalgal taxa. We propose a protocol for the quantitative assessment of heterogenous phytoplankton communities at the single-cell level and monitoring of phytoplankton bloom detection using a virtual filtering approach on a spectral flow cytometer (SFC-VF)., (© 2023. The Author(s).)

Spectral flow cytometry is a new technology that enables measurements of fluorescent spectra and light scattering properties in diverse cellular populations with high precision. Modern instruments allow simultaneous determination of up to 40+ fluorescent dyes with heavily overlapping emission spectra, discrimination of autofluorescent signals in the stained specimens, and detailed analysis of diverse autofluorescence of different cells-from mammalian to chlorophyll-containing cells like cyanobacteria. In this paper, we review the history, compare modern conventional and spectral flow cytometers, and discuss several applications of spectral flow cytometry., (© 2023. The Author(s).)