Sex preselection: High-speed flow cytometric sorting of X and Y sperm for maximum efficiency

Johnson, Lawrence A.; Welch, G. Rickey

doi:10.1016/s0093-691x(99)00220-4

Cited by 201 publications

(125 citation statements)

References 40 publications

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“…167 Further refinements to standard FC technology included replacement of the beveled needle by an orientating nozzle with a ceramic tip giving sperm less time to lose orientation, which improved correct orientation of sperm to 70%. 168 Conversion to highspeed modified FC operating under increased pressure had improved the accuracy and efficiency (20 000-25 000 events per second) of sperm sorting and purity of sorted population. 169 Although it was recently reported by some groups that stressful process of sorting might weaken status of livestock spermatozoa, strategies for restoring and strengthening their potentials were also discussed.…”

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

143

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%

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

Hossain¹,

Johannisson²,

Wallgren³

et al. 2011

Asian J Androl

143

120

View full text Add to dashboard Cite

show abstract

“…Before droplets disrupt from the discontinuous stream, the last hanging drop is charged according to the DNA content of the encased spermatozoon, which then passes an electrostatic field (3000 V), and droplets are deflected according to their charge. Sorted cells are pushed into a collection medium from where they are distributed to further preservation steps (Johnson & Welch 1999b). This set-up allows identification and selection of individual spermatozoa with sort purity above 90% of the desired characteristics (Johnson & Welch 1999b).…”

Section: Sex Sorting Of Spermatozoa By Flow Cytometrymentioning

confidence: 99%

Sex selection of sperm in farm animals: status report and developmental prospects

Rath¹,

Barcikowski²,

Graaf³

et al. 2013

Reproduction

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Pre-selection of spermatozoa based on the relative DNA difference between X-and Y-chromosome bearing populations by flow cytometry is an established method that has been introduced into commercial cattle production. Although several important improvements have increased the sort efficiency, the fertilising ability of sexed spermatozoa based on offspring per insemination is still behind farmers' expectations. The main stress factors, especially on mitochondria, that reduce the lifespan of spermatozoa are described, and new technical as well as biological solutions to maintain the natural sperm integrity and to increase the sorting efficiency are discussed. Among these methods are the identification of Y-chromosome bearing spermatozoa by bi-functionalised gold nanoparticles and triplex hybridisation in vivo as well as new laser-controlled deflection system that replaces the deflection of spermatozoa in the electrostatic field. Additionally, as well as a new nonsurgical transfer system of spermatozoa into the oviduct of cows has been developed and allows a significant reduction of spermatozoa per transfer. Altogether, the improvements made in the recent years will allow a broader use of sex-sorted spermatozoa even in those species that require more cells than cows and sheep.Reproduction (2013) 145 R15-R30

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“…Embora com bons resultados na separação espermática, a citometria de fluxo é pouco eficiente em relação ao investimento com o equipamento e pela baixa produção/ equipamento/hora, que não ultrapassa 12 × 10 6 espermatozoides em alta velocidade de separação (Johnson & Welch, 1999), restringindo sua utilização na indústria da inseminação artificial. Esse quadro é agravado pelo fato de que o sêmen de alguns deles não resiste ao processo de sexagem por essa técnica ou produzem variações na taxa de clivagem (0 a 89%) e na taxa de blastocistos produzidos por oócitos (3,5 a 28,8%) (Palma et al, 2008).…”

Section: Introductionunclassified

Sexagem de espermatozoides bovinos por centrifugação em gradiente descontínuo de densidade de Percoll

Lima¹,

Moreira‐Filho

Lucio

et al. 2011

R. Bras. Zootec.

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RESUMO -O objetivo neste trabalho foi desenvolver um método de seleção do sexo de espermatozoides bovinos por centrifugação em gradiente de densidade de Percoll. Utilizou-se sêmen congelado de touros mantidos em regime de colheita de sêmen. A fração de espermatozoides X ou Y foi separada por centrifugação em treze diferentes gradientes de densidade de Percoll formados por 1 a 12 camadas com densidades que variaram de 1,004 g/mL a 1,123 g/mL. As soluções com diferentes densidades foram preparadas misturando-se, em proporções diferentes, meio de cultura Hank's e uma solução estoque composta de NaCl 1,5 M e Percoll (1:9, v/v). Sobre cada gradiente foi colocado um total de 50 × 10 6 espermatozoides descongelados em 0,7 mL de meio Hank's e centrifugados a 250 X g por 30 minutos, em rotor horizontal, a 25°C. Os espermatozoides das frações superior e inferior foram tratados com Quinacrina Mustarda e analisados (200 deles) quanto à presença do corpúsculo-F. Dos espermatozoides encontrados no sedimento de dois gradientes, compostos de 8 e 12 frações com densidades variando entre 1,050 a 1,120 g/mL e 1,044 a 1,123 g/mL, respectivamente, visualizaram-se 25% com corpúsculo-F e os 75% restantes prováveis portadores do cromossomo X. O aumento na porcentagem de espermatozoides X após a centrifugação em gradiente de densidade permitirá que esse método de sexagem seja usado em larga escala na produção comercial de carne e leite bem como no teste de progênie. Palavras-chave: Bos taurus, seleção do sexo, sêmenSex selection in bovine spermatozoa by using Percoll discontinuos density gradient centrifugation ABSTRACT -The objective of this work was to develop a bovine spermatozoid sex selection method by using Percoll density gradient centrifugation. It was used frozen semen of bulls kept in semen collection regime. Fraction X or Y was separated by centrifugation in three different Percoll density gradient formed by 1 to 12 layers with densities varying from 1.004 g/mL to 1.123 g/mL. Solutions with different densities were prepared by mixing, at different proportions, Hank's culture medium and a stock solution composed of NaCl 1.5 M and Percoll (1:9, v/v). On each gradient, it was put 50 × 10 6 spermatozoids thawed in Hank's medium and centrifuged at 250 X g for 30 min in a horizontal rotor, at 25°C. The spermatozoids in the superior and inferior fractions were treated with Mustard Quinacrine and analyzed (200 spermatozoids) for the presence of F-body. Of the spermatozoids found in the sediment of the two gradients, composed of 8 and 12fractions with densities varying from 1.050 to 1.120 g/mL and 1.044 to 1.123 g/mL, respectively, it was identified 25%with the F-body and the other 75% were probably X chromosome-bearing spermatozoids. Increase in percentage of the X-spermatozoid after density gradient centrifugation will allow this system of spermatozoid sexing to be used in large scale in commercial production of meat and milk as well as for progeny tests in bovines. IntroduçãoA sexagem de espermatozoides tem valor econômico signifi...

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Sex preselection: High-speed flow cytometric sorting of X and Y sperm for maximum efficiency

Cited by 201 publications

References 40 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

Sex selection of sperm in farm animals: status report and developmental prospects

Sexagem de espermatozoides bovinos por centrifugação em gradiente descontínuo de densidade de Percoll

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