A highly efficient method for washing mammalian spermatozoa

Harrison, R. A. P.

doi:10.1530/jrf.0.0480347

Cited by 105 publications

(55 citation statements)

References 14 publications

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“…After dilution in 0-1 M-phosphate-buffered saline (PBS, pH 6-9) the spermatozoa were washed by centrifugation through 7-5% (w/v) Ficoll 400 (Pharmacia, Uppsala, Sweden) in PBS to remove seminal plasma components (Harrison, 1976 (Foulkes, 1977). The eluate volume was reduced with an Amicon CH4 concentrator and the protein concentration was determined (Lowry, Rosebrough, Farr & Randall, 1951).…”

Section: Preparation Ofspermatozoamentioning

confidence: 99%

A spectrofluorometric investigation, using 1-anilino-naphthalene-8-sulphonate, of the interaction between washed bovine spermatozoa and seminal plasma or egg-yolk lipoprotein

MacDonald¹,

Foulkes²

1981

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Summary. The dye 1-anilino-naphthalene-8-sulphonate (ANS) bound at 94\m=.\5\ m=+-\ 2\m=.\7 pmol per 106 spermatozoa (Kd = 1\m=.\83\ m=+-\0\m=.\13\m=x\ 10\m=-\5 m) and 7\m=.\30\ m=+-\ 0\m=.\17 pmol per mg seminal plasma protein (Kd = 2\m=.\19\ m=+-\0\m=.\06\m=x\10\m=-\5 m). Equilibration of excess ANS with a mixture of washed spermatozoa and seminal plasma resulted in a significant increase in fluorescence over that calculated by summation of the individual levels (P < 0\m=.\05). A less pronounced but significant increase was seen when an egg-yolk lipoprotein, previously shown to have cryoprotective properties, was similarly added to washed spermatozoa (P < 0\m=.\05).This increase in fluorescence was not reduced by washing spermatozoa\p=n-\lipoproteinmixtures, suggesting that an interaction between the lipoprotein and the cell membrane had occurred.

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Section: Preparation Ofspermatozoamentioning

confidence: 99%

A spectrofluorometric investigation, using 1-anilino-naphthalene-8-sulphonate, of the interaction between washed bovine spermatozoa and seminal plasma or egg-yolk lipoprotein

MacDonald¹,

Foulkes²

1981

Reproduction

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“…Harrison (1976) found that the technique of washing by centrifuging through Ficoll was highly efficient. In thisstudywe determined the effect on freezing damage after washing the spermatozoa upto4 times instead of theusual 2 times, andfound noimprovement.…”

Section: Resultsmentioning

confidence: 99%

“…Washing of spermatozoa Spermatozoa werewashedfreeof seminal plasmaby centrifugation through a solution ofFicoll(7%w/vin diluent) asdescribed by Harrison (1976). Thewashed spermatozoa werere-suspended indiluent andwashed a second time.…”

Section: Semen Diluentmentioning

confidence: 99%

Deleterious effect of seminal plasma in the cryo-preservation of bovine spermatozoa

Martinus

Molan

Shannon

1991

New Zealand Journal of Agricultural Research

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The effect of seminal plasma on spermatozoa wasstudied duringfreezing inabuffered diluent. The effect was assessed after thawing, both by visualassessment of motility and by colorimetric assayof aromatic L-aminoacid oxidase activity. The presence of seminal plasma caused damage to the spermatozoa, as reflected byan increase in therelease of the amino acid oxidase (increased from 39.6 to 86.1% of that released from maximally damaged spermatozoa) and a reduction in the motility of the spermatozoa (reduced from 29.6 to 2.7% motility). Verylittleof the damagewas incurred in thecooling of the spermatozoa to 5°C beforefreezing: this tends to suggest that the effect of seminal plasma on spermatozoa during freezing isdistinct from itsknown effectin cold shockof spermatozoa.

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“…Moreover, high concentrations of glycerol can overcome the inhibitory effect of a-chlorohydrin on ram spermatozoa (Edwards et al, 1976;Brown-Woodman et al, 1978), suggesting competition between the two compounds for the same pathway. Such competition is supported by the report that a-chlorohydrin is a competitive inhibitor of glycerol kinase prepared from Escherichia coli (Thorner, 1972; see Brooks, Hamilton & Mallek, 1974 (Blackshaw, 1954) and the spermatozoa were separated from seminal plasma by the procedure of Harrison (1976). The pellet of spermatozoa was frozen and subsequently thawed and sonicated for a total of 1 min in 50 mM-triethanolamine, 1 iriM-EDTA, 2 mM-MgCl2, 30 mM-mercaptoethanol, 0-75% Triton ¬ 00, pH 7-5.…”

Section: Introductionmentioning

confidence: 88%

The interaction of -chlorohydrin with glycerol kinase

De¹

1979

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A highly efficient method for washing mammalian spermatozoa

Cited by 105 publications

References 14 publications

A spectrofluorometric investigation, using 1-anilino-naphthalene-8-sulphonate, of the interaction between washed bovine spermatozoa and seminal plasma or egg-yolk lipoprotein

A spectrofluorometric investigation, using 1-anilino-naphthalene-8-sulphonate, of the interaction between washed bovine spermatozoa and seminal plasma or egg-yolk lipoprotein

Deleterious effect of seminal plasma in the cryo-preservation of bovine spermatozoa

The interaction of -chlorohydrin with glycerol kinase

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