Probes and Techniques for Sperm Evaluation by Flow Cytometry

Martínez‐Pastor, Felipe; Mata-Campuzano, M.; Álvarez‐Rodríguez, Manuel; Álvarez, M.; Anel, L.; Paz, P. de

doi:10.1111/j.1439-0531.2010.01622.x

Cited by 154 publications

(166 citation statements)

References 112 publications

(227 reference statements)

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“…149 This dye can be used together with viability dyes for removing the dead spermatozoa. 150 Dihydroethidium (hydroethidine) may be used as a specific probe to detect superoxide anion ( Figure 9); it is freely permeable and oxidized when bound to DNA, thus becoming valuable to detect superoxide anions in spermatozoa; 151 however, other oxygen species might also be detected, depending on the filters used for detection. 150 The MitoSOX (Invitrogen, Carlsbad, CA, USA) is another new probe able to detect mitochondrial superoxide in the live-cell population of human spermatozoa.…”

Section: Detection Of Oxidative Stress and Lipid Peroxidationmentioning

confidence: 99%

“…150 Dihydroethidium (hydroethidine) may be used as a specific probe to detect superoxide anion ( Figure 9); it is freely permeable and oxidized when bound to DNA, thus becoming valuable to detect superoxide anions in spermatozoa; 151 however, other oxygen species might also be detected, depending on the filters used for detection. 150 The MitoSOX (Invitrogen, Carlsbad, CA, USA) is another new probe able to detect mitochondrial superoxide in the live-cell population of human spermatozoa. 152 The probe is readily oxidized by superoxide, while prevented by superoxide dismutase, thus enabling highly fluorescent oxidation products to be detectable upon binding to nucleic acid.…”

Section: Detection Of Oxidative Stress and Lipid Peroxidationmentioning

confidence: 99%

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Flow cytometry for the assessment of animal sperm integrity and functionality: state of the art

Hossain¹,

Johannisson²,

Wallgren³

et al. 2011

Asian J Androl

135

120

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Flow cytometry is now a recognized methodology within animal spermatology, and has moved from being a research tool to become routine in the assessment of animal semen destined to breeding. The availability of 'bench-top' flow cytometers and of newer and versatile markers for cell structure and function had allowed the instrumentation to measure more sperm parameters, from viability to reactiveness when exposed to exogenous stimuli, and to increase our capabilities to sort spermatozoa for potential fertilizing capacity, or chromosomal sex. The present review summarizes the state of the art regarding flow cytometry applied to animal andrology, albeit keeping an open comparative intent. It critically evaluates the present and future capabilities of flow cytometry for the diagnostics of potential fertility and for the development of current reproductive technologies such as sperm freezing, sperm selection and sperm sorting. The flow cytometry methods will probably further revolutionize our understanding of the sperm physiology and their functionality, and will undoubtedly extend its application in isolating many uncharacterized features of spermatozoa. However, continuous follow-up of the methods is a necessity owing to technical developments and the complexity of mapping spermatozoa.

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Section: Detection Of Oxidative Stress and Lipid Peroxidationmentioning

confidence: 99%

Section: Detection Of Oxidative Stress and Lipid Peroxidationmentioning

confidence: 99%

Flow cytometry for the assessment of animal sperm integrity and functionality: state of the art

Hossain¹,

Johannisson²,

Wallgren³

et al. 2011

Asian J Androl

135

120

View full text Add to dashboard Cite

show abstract

“…Although many cytometric applications are now being adapted in veterinary andrology (51), some are redundant as they characterize essentially either the same functional properties as other applications or properties that are dependent upon other functions. Moreover, some tests can be performed by other means, without using cytometry.…”

Section: Cytometric Sperm Evaluationmentioning

confidence: 99%

“…Electronic cell counting, although it gives a good physical distribution of cells based on their size, has the obvious disadvantage that it is not possible to distinguish spermatozoa from other cells or other alien particles present in the ejaculate if these latter are of similar size. Flow cytometry encounters a similar problem if light scatter is used to select the population representing spermatozoa and needs additional tools (counting beads or livedead staining; (51,54). It has been demonstrated recently that this procedure leads to the systematic misestimation of cell counts.…”

Section: Sperm Concentrationmentioning

confidence: 99%

Cytometric solutions in veterinary andrology: Developments, advantages, and limitations

Petrunkina

Harrison²

2011

Cytometry Pt A

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Cytometric methodologies are becoming increasingly important in veterinary andrology as means of assessing sperm function. However, as yet, flow cytometric techniques in veterinary andrology have not kept up in sophistication with those in other areas of biology and medicine. In this brief review, we consider the present state of cytometry in andrological procedures for evaluating the fertility of domestic animal sires. We outline the aspects of sperm physiology, paying particular attention to the changes that take place during the process known as capacitation, which prepares the sperm for interaction with the egg. We then examine briefly but critically the cytometric techniques that are currently in commercial use or are being established in research laboratories for testing sperm characteristics. Current limitations and potential developments in semen assessment are discussed. Recent research knowledge offers possibilities for applying more subtle flow cytometric approaches to distinguish different levels of fertilizing potential in semen samples. For example, linking field fertility data to multiparametric kinetic studies of sperm capacitational changes rather than ''single parameter-single time point'' estimations may reveal that slower rather than rapid changes indicate high fertility. Moreover, the development of multicolor flow cytometric procedures as a means of evaluating multiple functional parameters in individual cells would reduce the uncertainties always inherent in predicting fertility from in vitro sperm evaluation tests. ' 2011 International Society for Advancement of Cytometry Key terms fertility; sperm evaluation; heterogeneity; subpopulations; multicolour cytometry THE main focus of an andrological assessment in veterinary medicine is to ascertain the fertility of a potential sire. Although the most obvious and indeed frequently used general veterinary test is to quantify the number of offspring born to the sire, this is a lengthy and very expensive process. For ''accuracy,'' many inseminations must be carried out, and results must await gestation and birth. Obviously, therefore, development of rapid tests that can be carried out in the laboratory has for long been important. However, as research into the reproductive process has advanced, it has become clear that the life of the fertilizing sperm between its release from the testis and its fusion with the egg is far from simple. Moreover, the population of sperm ejaculated into the female has been revealed as being heterogeneous in functional ability, even if morphologically normal [c.f. reviews (1,2)]. As a result of these findings, tests are being developed to examine functional ability within a heterogeneously responding population (3-5).Given that a normal male ejaculate contains many millions of individual sperm cells, it is natural that these tests are based on cytometry. In this short review, we will outline and comment upon a range of modern and still-developing cytometric methodologies in use or potentially useful for sperm...

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“…quality of spermatozoa (3). Acrosome and plasma membrane integrity are two sperm parameters often used in routine quality analysis of spermatozoa.…”

mentioning

confidence: 99%

Comparison of electronic volume and forward scatter principles of cell selection using flow cytometry for the evaluation of acrosome and plasma membrane integrity of bull spermatozoa

Standerholen

Myromslien²,

Kommisrud³

et al. 2014

Cytometry Pt A

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The objective of the study was to compare two different flow cytometers to reveal if there are differences between them and to find the most suitable protocol for analysis of spermatozoa. These two flow cytometers; Cell Lab Quanta TM and Coulter Epics XL, have different principles to calculate cell size, electric volume, and forward scatter (FS), respectively. Flow cytometry is a valuable tool to assess various spermatozoa quality traits simultaneously, such as plasma membrane and acrosome integrity. A double-and triple-stain combination was performed to compare evaluation of these two parameters by both flow cytometers and to assess the need of a fluorescent probe to identify the spermatozoa. Propidium iodide was used to assess the proportion of dead spermatozoa, whereas Alexa Fluor V R 488 conjugated peanut agglutinin (PNA-Alexa 488) was used to evaluate the percentage of acrosome intact and acrosome-reacted cells or degenerated cells. In the triple-stain protocol, MitoTracker V R Orange (MO) was included to test the capacity of this probe to discriminate spermatozoa from egg yolk and debris particles present in the semen sample. Cryopreserved semen from 13 Norwegian Red bulls was included in the study and the semen was evaluated immediately after thawing and after 3 hr incubation at 37 C. The results show that there is good agreement between the instruments. Nevertheless, a significant difference was found in percentages of acrosome intact live spermatozoa (% AIL) when including MO as a spermatozoa identification probe, compared to assessment without MO, with the Coulter Epics XL, while no significant difference was found when including the probe with the Cell Lab Quanta. In conclusion, the results show that cell size measurement based on electronic volume used by the Cell Lab Quanta flow cytometer is more accurate than FS used by the Coulter Epics XL flow cytometer in identification of spermatozoa. V C 2014 International Society for Advancement of Cytometry Key terms flow cytometry; sperm analysis; electronic volume; forward scatter; plasma membrane integrity; acrosome integrity FLOW cytometry has become a powerful tool for characterization of cell populations. The instrument has the capability to measure multiple parameters of individual cells, thus it is possible to detect abnormal cells in a short time (1-3). The flow cytometric analysis is based on light-spreading and fluorescence signals. Light that is scattered at high angles is termed side scatter (SS) and will provide information about such intracellular properties as cell granularity (4). Scattered light with a small angle is termed forward scatter (FS) and depends on particle size and the refractive

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Probes and Techniques for Sperm Evaluation by Flow Cytometry

Cited by 154 publications

References 112 publications

Flow cytometry for the assessment of animal sperm integrity and functionality: state of the art

Flow cytometry for the assessment of animal sperm integrity and functionality: state of the art

Cytometric solutions in veterinary andrology: Developments, advantages, and limitations

Comparison of electronic volume and forward scatter principles of cell selection using flow cytometry for the evaluation of acrosome and plasma membrane integrity of bull spermatozoa

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